U.S. patent application number 17/382945 was filed with the patent office on 2022-01-27 for methods for nomination of nuclease on-/off-target editing locations, designated "ctl-seq" (crispr tag linear-seq).
The applicant listed for this patent is INTEGRATED DNA TECHNOLOGIES, INC.. Invention is credited to Ellen BLACK, Keith GUNDERSON, Kyle KINNEY, Matthew MCNEILL, Garrett RETTIG, Chris SAILOR, Yongming SUN, Rolf TURK, Yu WANG.
Application Number | 20220025365 17/382945 |
Document ID | / |
Family ID | |
Filed Date | 2022-01-27 |
United States Patent
Application |
20220025365 |
Kind Code |
A1 |
MCNEILL; Matthew ; et
al. |
January 27, 2022 |
METHODS FOR NOMINATION OF NUCLEASE ON-/OFF-TARGET EDITING
LOCATIONS, DESIGNATED "CTL-seq" (CRISPR Tag Linear-seq)
Abstract
Described herein are methods for identifying and nominating on-
and off-target CRISPR editing sites with improved accuracy and
sensitivity.
Inventors: |
MCNEILL; Matthew; (Iowa
City, IA) ; TURK; Rolf; (Iowa City, IA) ;
RETTIG; Garrett; (Coralville, IA) ; BLACK; Ellen;
(Swisher, IA) ; SUN; Yongming; (San Ramon, CA)
; SAILOR; Chris; (Cedar Rapids, IA) ; WANG;
Yu; (North Grafton, MA) ; GUNDERSON; Keith;
(Iowa City, IA) ; KINNEY; Kyle; (Iowa City,
IA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
INTEGRATED DNA TECHNOLOGIES, INC. |
Coralville |
IA |
US |
|
|
Appl. No.: |
17/382945 |
Filed: |
July 22, 2021 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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63055460 |
Jul 23, 2020 |
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International
Class: |
C12N 15/11 20060101
C12N015/11; C12N 9/22 20060101 C12N009/22; C12Q 1/6853 20060101
C12Q001/6853 |
Claims
1. A method for identifying and nominating on- and off-target
CRISPR edited sites with improved accuracy and sensitivity, the
process comprising the steps of: (a) co-delivering a guide sequence
RNA (sgRNA) or a two-part CRISPR RNA:trans-activating crRNA
(crRNA:tracrRNA) duplex, one or more tag sequences, and an
RNA-guided endonuclease to cells; (b) incubating the cells for a
period of time sufficient for double strand breaks to occur; (c)
isolating genomic DNA from the cells, fragmenting the genomic DNA,
and ligating the fragmented genomic DNA to a unique molecular index
containing a universal adapter sequence; (d) amplifying the ligated
DNA fragments using primers targeting the tag and universal adapter
sequences to produce a first set of amplified sequences; (e)
amplifying the first set of amplified sequences using universal
sequencing primers targeting the tails of Tag-pTOP or Tag-pBOT
primers to produce a second set of amplified sequences; (f)
sequencing the pooled sequences and obtaining sequencing data; and
(g) identifying on-/off-target CRISPR editing loci.
2. The method of claim 1, wherein the universal sequencing primers
target SP1 or SP2 sequence (SEQ ID NO: 7, 8) tails on the Tag-pTOP
or Tag-pBOT primers to produce a second set of amplified
sequences.
3. The method of claim 1, wherein the universal sequencing primers
target predesigned non-homologous sequence (SEQ ID NO: 269-273)
tails on the Tag-pTOP or Tag-pBot primers to produce a second set
of amplified sequences.
4. The method of claim 1, wherein the universal sequencing primers
target predesigned 13-mer tails on the Tag-pTOP or Tag-pBot primers
to produce a second set of amplified sequences.
5. The method of claim 1, wherein step (g) comprises executing on a
processor: aligning the sequence data to a reference genome; (ii)
identifying on-/off-target CRISPR editing loci; and (iii)
outputting the alignment, analysis, and results data as
custom-formatted files, tables or graphics.
6. The method of claim 1, further comprising a step following step
(e) comprising: (e1) normalizing the second set of amplified
sequences to produce concentration normalized libraries, pooling
the normalized libraries with other samples to produce pooled
libraries; and continuing with steps (f)-(i).
7. The method of claim 1, wherein step (d) uses a supression PCR
method.
8. The method of claim 1, wherein the RNA-guided endonuclease
comprises an endogenously-expressed Cas enzyme, a Cas expression
vector, a Cas protein, or a Cas RNP complex.
9. The method of claim 1, wherein the RNA-guided endonuclease
comprises an endogenously-expressed Cas9 enzyme, a Cas9 expression
vector, a Cas9 protein, or a Cas9 RNP complex.
10. The method of claim 1, wherein the cells comprise human or
mouse cells.
11. The method of claim 1, wherein the period of time is about 24
hours to about 96 hours.
12. The method of claim 1, wherein multiple tag sequences are
co-delivered.
13. The method of claim 1, wherein the tag sequences comprise
double-stranded deoxyribooligonucleotides (dsDNA) comprising
52-base pairs.
14. The method of claim 1, wherein the tag sequences comprise a
5'-terminal phosphate, and phosphorothioate linkages between the
1.sup.st and 2.sup.nd, 2.sup.nd and 3.sup.rd, 50.sup.th and
51.sup.st, and 51.sup.st and 52.sup.nd nucleotides.
15. The method of claim 1, wherein the tag sequences comprise a
double stranded DNA comprising the complementary top and bottom
strand pairs of SEQ ID NO: 1-2 or 7-268.
16. On- and off-target CRISPR editing sites identified or nominated
using the method of claim 1.
17. A method for designing 52-base pair tag sequences, the method
comprising, executing on a processor: (a) randomly generating
13-nucleotide sequences with 40-90% GC content, max homopolymer
length A:2, C:3, G:2, T:2, weighted homopolymer rate <20,
self-folding T.sub.m<50.degree. C., and self-dimer
T.sub.m<50.degree. C.; (b) removing sequences that perfectly
align to a particular genome or that are homopolymers or GG or CC
dinucleotide motifs and obtaining a set of 13-mers; (c) selecting a
subset of the 13-mer sequences that contain one or less CC or GG
dinucleotide motifs; (d) concatenating four of the of 13-mer subset
sequences to form random 52-mer sequences; (e) aligning the random
52-mer sequences to a genome; (f) removing the random 52-mer
sequences that have similarity to the genome to produce a subset of
52-mer sequences; and (h) outputting the subset of 52-mer sequences
and generating the complementary strands to produce double stranded
52-base pair tag sequences.
18. The method of claim 17, wherein the genome is human or
mouse.
19. The method of claim 17, wherein the 52-base pair tag sequences
are-non complementary to the genome.
20. The method of claim 17, further comprising designing primers
for the 52-base pair tag sequences.
21. The method of claim 17, wherein the 52-base pair tag sequences
comprise a 5'-terminal phosphate, and phosphorothioate linkages
between the 1.sup.st and 2.sup.nd, 2.sup.nd and 3.sup.rd, 50.sup.th
and 51.sup.st, and 51.sup.st and 52.sup.nd nucleotides of the
52-base pair tag sequences.
22. The method of claim 17, further comprising synthesizing
oligonucleotides comprising the 52-base pair tag sequences, the
complement of the 52-base pair tag sequences, or primers for the
52-base pair tag sequences.
23. One or more 52-base pair tag sequences designed using the
methods of claim 17.
24. The 52-base pair tag sequences of claim 23, wherein the 52-base
pair tag sequence comprises a double stranded DNA comprising the
top and bottom strand pairs of SEQ ID NO: 1-2 or 7-268.
25. A method for designing primers partially complementary to the
52-base pair tag sequences of claim 23 and an adapter primer, the
method comprising, executing on a processor: (a) designing tag
primers that are partially complementary to the top and bottom
strands of tag sequences; and (b) designing an adapter primer that
is partially complementary to the top strand of the adapter
sequence; wherein: the tag primers comprise a 5'-universal tail
sequence; and the adapter primer comprises a sequence complementary
to the tails of Tag-pTOP or Tag-pBOT primers.
26. The method of claim 25, wherein the 5'-universal tail sequence
is complementary to an SP1 or SP2 sequence (SEQ ID NO: 7, 8), a
locus specific segment, a ribonucleotide (rN) 6-nucleotides from
the 3'-end, a 3'-end mismatch, a 3'-end block (3'-C.sub.3 spacer),
a predesigned non-homologous sequence (SEQ ID NO: 269-273), or a
predesigned 13-mer sequence.
27. The method of claim 25, wherein the primers partially
complementary to top and bottom strands of the tag sequences
comprise a tail sequence complementary to the SP1 sequence (SEQ ID
NO: 7) and the adapter primer comprises a sequence complementary to
the SP2 sequence (SEQ ID NO: 8) tail on the Tag-pTOP or Tag-pBOT
primers; or the primers partially complementary to top and bottom
strands of the tag sequences comprise a tail sequence complementary
to the SP2 sequence (SEQ ID NO: 8) and the adapter primer comprises
a sequence complementary to the SP1 sequence (SEQ ID NO: 7) tail on
the Tag-pTOP or Tag-pBOT primers.
28. The method of claim 25, wherein the amplification of a nucleic
acid molecule with the primers that are complementary to the top
and bottom strands of tag sequences and primers that are
complementary to the top strand of the adapter sequence produces a
PCR product that comprises a portion of the tag sequence, a sgDNA
sequence, and the adapter sequence.
29. The method of claim 25, further comprising synthesizing
oligonucleotides comprising the sequences of the forward and
reverse tag primers and the adapter primer.
30. The method of claim 25, wherein the 52-base pair tag sequences
and primers partially complementary to the 52-base pair tag
sequences are designed and selected using an algorithm predicting
whether the primers are likely to be partially complementary and
have a propensity to form primer-dimers.
31. One or more primers partially complementary to the 52-base pair
tag sequences and one or more adapter primers designed using the
method of claim 25.
32. The primers of claim 31, wherein the primers comprise the
sequences of SEQ ID NO: 3, 4; and the adapter primer, wherein the
adapter primer comprises the sequence of SEQ ID NO: 5.
33. A method for using of one or more double-stranded 52-base pair
tag sequences to identify on- and off-target CRISPR editing sites.
Description
CROSS REFERENCE TO RELATED APPLICATIONS
[0001] This application claims priority to U.S. Provisional Patent
Application No. 63/055,460, filed on Jul. 23, 2020, which is
incorporated by reference herein in its entirety.
REFERENCE TO SEQUENCE LISTING
[0002] This application is filed with a Computer Readable Form of a
Sequence Listing in accordance with 37 C.F.R. .sctn. 1.821(c). The
text file submitted by EFS,
"013670-9056-US02_sequence_listing_19-JUL-2021_ST25.txt" contains
273 sequences, was created on Jul. 19, 2021, has a file size of 153
Kbytes, and is hereby incorporated by reference in its
entirety.
TECHNICAL FIELD
[0003] Described herein are methods for identifying and nominating
on- and off-target CRISPR editing sites with improved accuracy and
sensitivity.
BACKGROUND
[0004] CRISPR (clustered regularly interspaced short palindromic
repeats) has revolutionized genomics by permitting the simple
introduction of changes to the genetic code. CRISPR systems, such
as Cas9 and Cas12a proteins, are guided to their target by RNA
oligonucleotide sequences bound by the Cas proteins (forming
ribonucleoprotein protein; RNP), where the enzyme creates double
stranded breaks (DSBs) in DNA sequences. Native cellular machinery
repairs DSBs, generally using non-homologous end joining (NHEJ) or
homology directed repair (HDR) molecular pathways. DNA repaired
through NHEJ, which occurs at on- and off-target locations, often
contains indels (insertions/deletions), which can lead to mutations
and change the function of encoded genes. Thus, identifying these
locations is critical to deconvoluting the impact of on- and
off-target editing on biological phenotypes.
[0005] To date, no "gold standard" method exists to identify or
nominate off-target editing locations for CRISPR or other
nucleases. Many methods have been developed. These methods use a
variety of strategies, including the detection of endogenous repair
machinery assembled at DSBs (Discover-Seq [1]), the integration of
a DNA tag sequence into the host cell genome (GUIDE-Seq; see U.S.
Pat. No. 9,822,407), iGUIDE [2, 3]), or by cutting DNA in vitro
(BLISS [4], CIRCLE-Seq [5], SiteSeq [6]).
[0006] Cellular or cell based (sometimes referred to as in vivo)
and biochemical (sometimes referred to as in vitro) off-target
assay nomination systems each have their advantages. Proteins bound
to the DNA and epigenetic marks modify the function of nuclease
activity, suggesting that cellular or cell based methods may better
identify actual editing targets [7]. However, biochemical methods
have nominated sites not identified through cellular or cell based
methods, suggesting biochemical methods may be more comprehensive
[5, 6]. Nevertheless, these current tools tend to have imperfect
sensitivity [5, 6] (see FIG. 1).
[0007] What is needed is a method for detecting and nominating on-
and off-target CRISPR editing sites with improved accuracy and
sensitivity.
SUMMARY
[0008] One embodiment described herein is a method for identifying
and nominating on- and off-target CRISPR edited sites with improved
accuracy and sensitivity, the process comprising the steps of: (a)
co-delivering a guide sequence RNA (sgRNA) or a two-part CRISPR
RNA:trans-activating crRNA (crRNA:tracrRNA) duplex, one or more tag
sequences, and an RNA-guided endonuclease to cells; (b) incubating
the cells for a period of time sufficient for double strand breaks
to occur; (c) isolating genomic DNA from the cells, fragmenting the
genomic DNA, and ligating the fragmented genomic DNA to a unique
molecular index containing a universal adapter sequence; (d)
amplifying the ligated DNA fragments using primers targeting the
tag and universal adapter sequences to produce a first set of
amplified sequences; (e) amplifying the first set of amplified
sequences using universal sequencing primers targeting the tails of
Tag-pTOP or Tag-pBOT primers to produce a second set of amplified
sequences; (f) sequencing the pooled sequences and obtaining
sequencing data; and (g) identifying on-/off-target CRISPR editing
loci. In one aspect, the universal sequencing primers target SP1 or
SP2 sequence (SEQ ID NO: 7, 8) tails on the Tag-pTOP or Tag-pBOT
primers to produce a second set of amplified sequences. In another
aspect, the universal sequencing primers target predesigned
non-homologous sequence (SEQ ID NO: 269-273) tails on the Tag-pTOP
or Tag-pBot primers to produce a second set of amplified sequences.
In another aspect, the universal sequencing primers target
predesigned 13-mer tails on the Tag-pTOP or Tag-pBot primers to
produce a second set of amplified sequences. In another aspect,
step (g) comprises executing on a processor: (i) aligning the
sequence data to a reference genome; (ii) identifying
on-/off-target CRISPR editing loci; and (iii) outputting the
alignment, analysis, and results data as custom-formatted files,
tables or graphics. In another aspect, the method further comprises
a step following step (e) comprising: (e1) normalizing the second
set of amplified sequences to produce concentration normalized
libraries, pooling the normalized libraries with other samples to
produce pooled libraries; and continuing with steps (f)-(i). In
another aspect, step (d) uses a suppression PCR method. In another
aspect, the RNA-guided endonuclease comprises an
endogenously-expressed Cas enzyme, a Cas expression vector, a Cas
protein, or a Cas RNP complex. In another aspect, the RNA-guided
endonuclease comprises an endogenously-expressed Cas9 enzyme, a
Cas9 expression vector, a Cas9 protein, or a Cas9 RNP complex. In
another aspect, the cells comprise human or mouse cells. In another
aspect, the period of time is about 24 hours to about 96 hours. In
another aspect, multiple tag sequences are co-delivered. In another
aspect, the tag sequences comprise double-stranded
deoxyribooligonucleotides (dsDNA) comprising 52-base pairs. In
another aspect, the tag sequences comprise a 5'-terminal phosphate,
and phosphorothioate linkages between the 1.sup.st and 2.sup.nd,
2.sup.nd and 3.sup.rd, 50.sup.th and 51.sup.st, and 51.sup.st and
52.sup.nd nucleotides. In another aspect, the tag sequences
comprise a double stranded DNA comprising the complementary top and
bottom strand pairs of SEQ ID NO: 1-2 or 7-268.
[0009] Other embodiments described herein are on- and off-target
CRISPR editing sites identified or nominated using the methods
described herein.
[0010] Another embodiment described herein is a method for
designing 52-base pair tag sequences, the method comprising,
executing on a processor: (a) randomly generating 13-nucleotide
sequences with 40-90% GC content, max homopolymer length A:2, C:3,
G:2, T:2, weighted homopolymer rate <20, self-folding
T.sub.m<50.degree. C., and self-dimer T.sub.m<50.degree. C.;
(b) removing sequences that perfectly align to a particular genome
or that are homopolymers or GG or CC dinucleotide motifs and
obtaining a set of 13-mers; (c) selecting a subset of the 13-mer
sequences that contain one or less CC or GG dinucleotide motifs;
(d) concatenating four of the of 13-mer subset sequences to form
random 52-mer sequences; (e) aligning the random 52-mer sequences
to a genome; (f) removing the random 52-mer sequences that have
similarity to the genome to produce a subset of 52-mer sequences;
and (h) outputting the subset of 52-mer sequences and generating
the complementary strands to produce double stranded 52-base pair
tag sequences. In one aspect, the genome is human or mouse. In
another aspect, the 52-base pair tag sequences are-non
complementary to the genome. In another aspect, the method further
comprises designing primers for the 52-base pair tag sequences. In
another aspect, the 52-base pair tag sequences comprise a
5'-terminal phosphate, and phosphorothioate linkages between the
1.sup.st and 2.sup.nd, 2.sup.nd and 3.sup.rd, 50.sup.th and
51.sup.st, and 51.sup.st and 52.sup.nd nucleotides of the 52-base
pair tag sequences. In another aspect, the method further comprises
synthesizing oligonucleotides comprising the 52-base pair tag
sequences, the complement of the 52-base pair tag sequences, or
primers for the 52-base pair tag sequences.
[0011] Other embodiments described herein are one or more 52-base
pair tag sequences designed using the methods described herein. In
one aspect, the 52-base pair tag sequence comprises a double
stranded DNA comprising the top and bottom strand pairs of SEQ ID
NO: 1-2 or 7-268.
[0012] Another embodiment described herein is a method for
designing primers partially complementary to the 52-base pair tag
sequences of claim 23 and an adapter primer, the method comprising,
executing on a processor: (a) designing tag primers that are
partially complementary to the top and bottom strands of tag
sequences; and (b) designing an adapter primer that is partially
complementary to the top strand of the adapter sequence; wherein:
the tag primers comprise a 5'-universal tail sequence; and the
adapter primer comprises a sequence complementary to the tails of
Tag-pTOP or Tag-pBOT primers. In one aspect, the 5'-universal tail
sequence is complementary to an SP1 or SP2 sequence (SEQ ID NO: 7,
8), a locus specific segment, a ribonucleotide (rN) 6-nucleotides
from the 3'-end, a 3'-end mismatch, a 3'-end block (3'-C.sub.3
spacer), a predesigned non-homologous sequence (SEQ ID NO:
269-273), or a predesigned 13-mer sequence. In another aspect, the
primers partially complementary to top and bottom strands of the
tag sequences comprise a tail sequence complementary to the SP1
sequence (SEQ ID NO: 7) and the adapter primer comprises a sequence
complementary to the SP2 sequence (SEQ ID NO: 8) tail on the
Tag-pTOP or Tag-pBOT primers; or the primers partially
complementary to top and bottom strands of the tag sequences
comprise a tail sequence complementary to the SP2 sequence (SEQ ID
NO: 8) and the adapter primer comprises a sequence complementary to
the SP1 sequence (SEQ ID NO: 7) tail on the Tag-pTOP or Tag-pBOT
primers. In another aspect, the amplification of a nucleic acid
molecule with the primers that are complementary to the top and
bottom strands of tag sequences and primers that are complementary
to the top strand of the adapter sequence produces a PCR product
that comprises a portion of the tag sequence, a sgDNA sequence, and
the adapter sequence. In another aspect, the method further
comprises synthesizing oligonucleotides comprising the sequences of
the forward and reverse tag primers and the adapter primer. In
another aspect, the 52-base pair tag sequences and primers
partially complementary to the 52-base pair tag sequences are
designed and selected using an algorithm predicting whether the
primers are likely to be partially complementary and have a
propensity to form primer-dimers.
[0013] Other embodiments described herein are one or more primers
partially complementary to the 52-base pair tag sequences and one
or more adapter primers designed using the methods described
herein. In one aspect, the primers comprise the sequences of SEQ ID
NO: 3, 4; and the adapter primer, wherein the adapter primer
comprises the sequence of SEQ ID NO: 5.
[0014] Another embodiment described herein is the use of one or
more double-stranded 52-base pair tag sequences for identifying on-
and off-target CRISPR editing sites.
DESCRIPTION OF THE DRAWINGS
[0015] FIG. 1 shows fraction of reads shared by three biological
replicates are shown in white sectors; whereas reads shared by two
replicates, or present in a single replicate, are shown in black
sectors. Table 1 shows GUIDE-seq [3] based nomination for 4
different gRNAs in triplicate in a 96-well format. gRNA complexes
were generated by mixing equimolar amounts of Alt-R crRNA-XT and
Alt-R tracrRNA. HEK293 cells stably expressing Cas9 were
transfected with 10 .mu.M gRNA and 0.5 .mu.M dsODN GUIDE-seq tag
using the Nucleofector.TM. system (Lonza). After 72 hrs, genomic
DNA (gDNA) was isolated. Genomic DNA was fragmented, and adapters
were ligated using the Lotus DNA library preparation kit (IDT).
Libraries were generated by amplification from the inserted tag to
the ligated adapters [3]. Libraries were then sequenced in
paired-end fashion on an IIlumina.RTM. platform.
[0016] FIG. 2 shows that GUIDE-Seq finds more off-target locations
than can be validated through rhAmpSeq targeted amplification.
Presented results are an aggregate of 331 GUIDE-Seq nominated sites
when delivering gRNA sequences (internally named: AR, CTNNB1, EMX1,
GRHPR, HPRT38087, HPRT38285, VEGFA) into HEK293 cells stably
expressing WT Cas9. GUIDE-seq nominated off-targets assigned 0.1%
of the total reference genome aligned reads for each guide were
designed and targeted by one rhAmpSeq panel all reference genome
aligned. In subsequent experiments, gRNAs were again delivered to
the same cells, and editing was assayed with rhAmpSeq. Targets were
called "edited" if the treated condition had observed indels
.gtoreq.the untreated control sample at %.
[0017] FIG. 3 illustrates that GUIDE-Seq tag integration rate
varies. The graph shows the percentage of Tag integration
(normalized to % Editing) for 118 unique Cas9 on/off-target sites
that had InDel editing in rhAmpSeq panels targeting GUIDE-Seq
nominated on/off-target loci for guide sequences targeting the
RAG1, RAG2, and EMX1 genes. Each guide was co-delivered with the
34-base pair GUIDE-Seq, dsODN tag into HEK293 cells stably
expressing Cas9 by nucleofection. DNA was extracted 72 hrs later,
amplified by rhAmpSeq multiplex PCR, sequenced on an Illumina.RTM.
MiSeq, and analyzed through a custom pipeline. The normalized tag
integration rate is calculated as the percentage of sequenced reads
at each target containing the tag sequence divided by the total
reads containing an allele divergent from the reference genome
(indicating Cas9 editing).
[0018] FIG. 4 shows the design of rhAmpSeq primers against alien
sequence tags. A cartoon diagram shows the steps of the design
process using the rhAmpSeq design pipeline including design of
forward primers against the top (1) and bottom (2) strands,
discarding unneeded primers, and selecting tag-targeting primers
that have 5'-overlapping, but not 3'-overlapping sequences, so that
the top/bottom strand primer dimers would hairpin (3).
[0019] FIG. 5 shows an overview of the rhAmpSeq design pipeline
used to construct the overlapping primer designs. In the pipeline,
a known sequence is appended onto the 5'-end and 3'-end of each tag
sequence, the inputs are quality-controlled and assays (shown in
FIG. 4A) are designed against the top and bottom strand of each
tag. Primers targeting each tag strand are paired such that at
least 4-nucleotides 3' of the RNA nucleotide do not overlap between
primers targeting the same tag, and primer pairs are ranked and
selected. Hg38 and mm38 acronyms represent versions of the human
and mouse genomes, respectively.
[0020] FIG. 6 illustrates hairpin formation if overlapping primers
generate PCR amplicons. The diagram shows a representative target
sequence and hairpin PCR product of undesired short amplicons from
overlapping primer regions with complementary 5' primer tail ends
at the 3'- and 5'-end of the PCR product.
[0021] FIG. 7 shows the number of target sites (black bars) with
integration of the specified single tag (SEQ ID NO: 9-40) or pools
of tags described in Table 5 (SEQ ID NO: 9-40, 45-268). The striped
bar (CTLmax) shows the maximum number of target sites that
theoretically can be found if a combination of the single tags (SEQ
ID NO: 9-40) is used (23 sites out of a maximum of 32 sites). Pool
A1 contains all the single tags (SEQ ID NO: 9-40). Pools B1-6
contain 16 different tags each (SEQ ID NO: 45-268). Pool C1
contains all tags tested (SEQ ID NO: 9-40, 45-268). Integration
events were determined using an in-house data analysis tool.
[0022] FIG. 8 shows the number of target sites (black bars) with
integration of the specified single tag (SEQ ID NO: 9-40) or pools
of tags described in Table 5 (SEQ ID NO: 9-40, 45-268). The striped
bar (CTLmax) shows the maximum number of target sites that
theoretically can be found if a combination of the single tags (SEQ
ID NO: 9-40) is used (47 sites out of a maximum of 53 sites). Pool
A1 contains all the single tags (SEQ ID NO: 9-40). Pools B1-6
contain 16 different tags each (SEQ ID NO: 45-268). Pool C1
contains all tags tested (SEQ ID NO: 9-40, 45-268). Integration
events were determined using an in-house data analysis tool.
DETAILED DESCRIPTION
[0023] Described herein are methods for detecting and nominating
on- and off-target CRISPR editing sites with improved accuracy and
sensitivity. The intracellular context information is maintained by
building upon prior in vivo nomination methods. The sensitivity is
expanded by co-delivering a set of unique, predefined sequence
tags. In one aspect, the co-delivered set of predefined unique tags
may range from 13-80 base pairs. In another aspect, the
co-delivered set of predefined tags may be comprised of 13 base
pair tag sequence tags, 26 base pair tag sequence tags, 39 base
pair tag sequence tags, 52 base pair tag sequence tags, 65 base
pair tag sequence tags, or 78 base pair tag sequence tags. In
another aspect, the unique predefined tags are a set of 52-base
pair tag sequence tags (the increased length of the sequence tags
improves the ability to find good primer landing sites for
rhPrimers). This limitation is believed to be mitigated by using a
diversity of tag sequences that are distinct from human and mouse
genomes. The specificity is improved by building upon Integrated
DNA Technologies (IDT)'s rhAmp technology that uses RNAaseH2
(Pyrococcus abyssi) to unblock primers that have correctly annealed
to their target; this yields lower rates of false priming.
Specificity can be further enhanced by only nominating targets
using reads that contain an expected tag sequence at the 5'-end.
The incorporation of suppression PCR into this method permits ease
of use. The prior in vivo methods (e.g., GUIDE-seq and iGUIDE)
require parallel PCR reactions (2 pool amplification) to amplify by
annealing to and extending from the top and bottom strand of the
tags. Here, suppression PCR is used to allow both pools to be
amplified simultaneously without causing problematic dimer
sequences.
[0024] A GUIDE-Seq dsDNA tag was co-delivered with one guide RNA to
HEK293 cells constitutively expressing Cas9 using nucleofection.
See U.S. Pat. No. 9,822,407, which is incorporated by reference
herein for such teachings. A total of four different guide RNAs
were tested in this fashion. Ribonucleoprotein complexes (RNPs)
between the expressed Cas9 and guide RNA form within the cells,
introducing double stranded breaks. Repaired breaks can contain the
co-delivered tags. After delivery, cells were incubated, and the
resulting DNA was extracted. Target amplification was performed
according to the GUIDE-Seq protocol and assayed with a modified
version of the GUIDE-Seq analytical pipeline
(github.com/aryeelab/guideseq). Nominated targets were compared
between three biological replicates (unique guideRNA+Tag
co-deliveries). Not all nominated targets were common to all
biological replicates (commonly/total nominated targets: 7/31,
6/19, 2/4, 3/5 respectively; see Table 1). However, >90% of the
total reads, attributed to any target, were attributed to common
targets (on average; see FIG. 1).
TABLE-US-00001 TABLE 1 Identified off-target sites for four
different gRNAs and relative level of editing at off-target sites
compared to the on-target site Location C19orf84_BR1 C19orf84_BR2
C19orf84_BR3 chr19_51389306 100.00% 100.00% 100.00% chr9_20224748
38.55% 16.43% 29.00% chr4_28036434 16.33% 13.05% 14.36%
chr15_74256506 14.30% 18.18% 25.17% chr2_171312919 11.40% 8.51%
7.93% chr8_65742269 10.82% 1.17% 10.40% chr13_96554656 8.70% 0.00%
0.00% chr4_86807920 8.50% 9.21% 1.92% chr3_124485356 6.57% 0.00%
0.00% chr9_20330398 5.60% 0.00% 0.00% chr11_71298123 5.12% 0.00%
0.00% chr7_101729696 4.83% 0.00% 9.58% chr19_10923882 3.67% 3.03%
0.00% chr10_15548456 3.57% 15.38% 0.00% chr12_117097457 2.80% 0.00%
2.60% chr22_33493900 2.13% 0.00% 4.79% chrX_149763439 2.13% 0.00%
3.83% chr17_7435217 1.93% 0.00% 0.55% chr12_26286721 1.74% 0.00%
5.06% chr16_49704848 1.26% 5.01% 7.11% chr12_51288216 1.06% 0.00%
0.00% chr12_56010621 0.87% 0.00% 0.00% chr13_29717148 0.48% 0.00%
0.00% chr1_3088065 0.29% 0.00% 0.00% chr15_73442915 0.19% 0.00%
0.55% chr10_118045968 0.19% 0.00% 0.00% chr14_102199972 0.00% 0.00%
0.68% chr18_56334679 0.00% 0.00% 2.33% chr21_36426137 0.00% 0.00%
2.19% chr5_139002763 0.00% 0.00% 3.83% chrX_58291642 0.00% 0.00%
3.83% Location C17orf99_BR1 C17orf99_BR2 C17orf99_BR3
chr17_78164110 100.00% 100.00% 100.00% chr22_24471716 15.00% 13.24%
10.86% chr10_101156881 6.22% 11.07% 9.79% chr3_170476431 5.86%
3.97% 4.57% chr17_17692965 4.94% 0.66% 8.62% chr15_73400031 3.93%
4.63% 5.73% chr19_15238775 0.00% 0.00% 2.56% chr2_18362316 0.00%
0.00% 1.59% chr2_171087784 0.00% 0.54% 0.84% chr22_19959968 0.00%
1.26% 0.19% chr22_32114104 0.00% 0.00% 4.06% chr4_129034015 0.00%
0.00% 0.33% chr5_61219030 0.00% 0.00% 0.33% chr5_66209615 0.00%
0.00% 1.86% chr7_69709389 0.00% 0.12% 2.75% chr7_158662844 0.00%
1.44% 5.27% chrX_9567397 0.00% 0.00% 0.23% chr19_55657073 0.00%
0.66% 0.00% chr22_43788032 0.00% 2.47% 0.00% Location C16orf90_BR1
C16orf90_BR2 C16orf90_BR3 chr16_3494817 100.00% 100.00% 100.00%
chr2_109189307 75.32% 4.27% 52.05% chr22_24586001 45.45% 0.00%
0.00% chr10_104736568 0.00% 0.00% 8.22% Location ATAD3C_BR1
ATAD3C_BR2 ATAD3C_BR3 chr1_1450685 100.00% 100.00% 100.00%
chr1_1503588 11.73% 10.07% 9.27% chr1_1516015 2.47% 1.86% 5.14%
chr19_32167960 26.34% 0.93% 0.00% chr2_111077960 0.00% 1.12%
0.00%
[0025] Additionally, nominated targets may not be replicable or
detectable using orthogonal methods. Using the GUIDE-Seq method,
the GUIDE-Seq DNA tag was co-delivered with each of 6 guides (each
tag is delivered with one guide RNA) to HEK293 cells constitutively
expressing Cas9 using nucleofection. rhAmpSeq multiplex amplicon
panels were designed to amplify the nominated targets, and we
quantified editing in biological replicates. Of the 331 targets
nominated by GUIDE-Seq, only 41 (12%) could be verified with
rhAmpSeq (see FIG. 2).
[0026] dsDNA tag sequences co-delivered with the guide RNAs into a
stably expressing CRISPR cell line, which are used in the NHEJ
repair, are incorporated at varying rates. Here, the GUIDE-Seq
dsDNA tag was co-delivered with each of 6 guides into HEK293 cells
constitutively expressing Cas9. In another aspect, the dsDNA tag
sequences co-delivered with CRISPR RNP, which are used in the NHEJ
repair, are incorporated at varying rates. Here, the GUIDE-Seq
dsDNA tag was co-delivered with each of 6 guides into HEK293 cells
constitutively expressing Cas9. rhAmpSeq panels were developed to
amplify nominated targets, and in biological replicates, the rates
of tag integration were analyzed using a custom analytical
pipeline. These results demonstrate that tags are incorporated at
0-85% of edited genomic copies, varying by target (see FIG. 3).
Without being bound by any theory, it is hypothesized that the rate
varies by sequence context.
[0027] Described herein are methods to improve the signal to noise
ratio by combining Integrated DNA Technology's rhAmpSeg.TM.
technology, suppression PCR, and novel alien DNA sequence designs
to nominate nuclease off-target editing locations within a host
genome.
[0028] In this method, Cas9, a sgRNA or a two-part CRISPR
RNA:trans-activating crRNA (crRNA:tracrRNA) duplex, and one or more
double stranded DNA (dsDNA) tag sequences are delivered to cells.
Co-delivering multiple tags permits improved tag integration at
off-target sites (see below). The tag sequences have sequence
content significantly different (i.e., alien) to the host genome.
After nuclease introduced DSBs, NHEJ repair will insert the tag
sequence(s) into the target site, forming known primer landing
sites. After cells have time to repair the DSBs and possibly
further divide (such as after 72 hr), genomic DNA is isolated,
fragmented (e.g., Covaris.RTM. shearing, enzyme-based shearing,
Tn5, etc.), ligated a unique molecular index (UMI)-containing
universal adapter sequence to the fragmented DNA, and the
un-ligated material is removed. Next, the DNA fragments are
amplified by targeting primers to the tag and universal adapter
sequences (Round 1 PCR). Using universal primers, a sample index
(PCR2) is added, the amplified material is concentration
normalized, pooled with other samples, and the pooled material is
sequenced on an IIlumina.RTM. (or similar) machine. The sequenced
reads are aligned to a reference genome, and loci where large
numbers of reads map may nominate on/off-target locations.
[0029] Alien sequences were designed by generating >1 M random
13-mer sequences with 40-90% GC content, max homopolymer length
A:2, C:3, G:2, T:2, weighted homopolymer rate <20, self-folding
T.sub.m<50.degree. C., and self-dimer T.sub.m<50.degree. C.
From the list of sequences, sequences that aligned perfectly
against human (GRCh38.p2; hg38) or mouse (GRCh38.p4; mm38)
reference genomes or had troubling motif sequences (homopolymers,
most G-G or C-C dinucleotide motifs) were removed, resulting in 479
sequences.
[0030] To design the 52-base pair tag sequences described herein,
49 13-mer oligo sequences were selected that contain .ltoreq.1 C or
G dinucleotide, and 10,000 unique combinations of four 13-mer
sequences were generated. The length of each concatenated sequence
(e.g., pasting four 13-mer sequences in a row using software) is
52-nucleotides. Next, each 52-nucleotide tag sequence was aligned
against the human (GRCh38.p2) and mouse (GRChm38.p4) genomes using
an internally modified version of bwa, called bwa-psm.
Implementation of bwa-psm returns all possible secondary matches up
to a defined threshold. A set of tag sequences (SEQ ID NO:1-2) were
designed that were intended to work as a group, that had no
similarity to the human or mouse genomes (max seed size: 7, seed
edit distance: 2, max edit distance: 21, max gap open: 2, max gap
extension: 3, mismatch penalty: 1, gap open penalty: 1, gap
extension penalty: 1).
[0031] Overlapping rhAmpSeq V1 primers (SEQ ID NO: 3-4) were
designed complementary to the top and bottom strands of the tag and
5'-end of the adapter sequence (SEQ ID NO: 6) (FIG. 4). The
tag-specific primers (SEQ ID NO: 3-4) contain a 5'-universal tail
sequence matching the SP1 and SP2 primer sequences (SEQ ID NO:
7-8), a locus specific segment, a ribonucleotide (rN) 6-nucleotides
from the 3'-end, a 3'-end mismatch, and a 3'-end block (3'-C.sub.3
spacer). The adapter-specific primer (SEQ ID NO: 5) targets the
5'-end of the 5'-P5 adapter sequence (SEQ ID NO: 6), and the
adapter sequence contains unique molecular index (UMI) sequence
(Table 2). The primers were designed to target the plus and minus
strands of the annealed tag such that, if these primers
unexpectedly form a dimer, the formed product will hairpin,
removing the oligo from the available reaction templates (e.g.,
supression PCR). (FIG. 6A-B). Primer sequences targeting the tags
were chosen based on a proprietary design algorithm designed and
implemented by IDT (internal copy of the algorithm with a
public-facing UI:
www.idtdna.com/site/account?RetumURL=/site/order/designtool/index/RHAMPSE-
Q), which selects the most optimally performing primer pairs to
amplify the intended template sequence. (FIG. 5). Primer sequences
were assessed for non-specific binding to all other tag sequences
and both human and mouse primary genome assemblies to verify they
were unlikely to form off-target amplicons when combined with a
universal adapter sequence and the presence of human or mouse
genomic DNA.
[0032] The primers were desired to work in pairs where one
tag-specific primer (top or bottom strand) pairs with the
adapter-specific primer (SEQ ID NO:5). This results in the
amplification of a molecule that contains a portion of the tag,
gDNA, and the adapter sequence when amplified using supression PCR
methods (FIG. 4).
TABLE-US-00002 TABLE 2 Sequences Used for First Proof of Concept
SEQ Sequence ID Type Name (5'.fwdarw.3') NO Tag 9022179029169042579
T*C*GTTCGTTC SEQ 04625907201907281 CGCTCTAACCGG ID CGAATCTACCGC NO:
GCATATCTACGC 1 CGCA*A*T Tag 9022179029169042579 A*T*TGCGGCGT SEQ
04625907201907281_r AGATATGCGCGG ID ev TAGATTCGCCGG NO:
TTAGAGCGGAAC 2 GAAC*G*A Tag pFWD.ID_Target1: acactctttccc SEQ
Primers 9022179029169042579 tacacgacgctc ID 04625907201907281.12
ttccgatctTCT NO: 7.150.1.SP1 ACCGCGCATATC 3 TACrGCCGCT/ 3SpC3/ Tag
pFWD.ID_Target2: acactctttccc SEQ Primers 9022179029169042579
tacacgacgctc ID 04625907201907281.11 ttccgatctATA NO: 6.140.-1.SP1
TGCGCGGTAGAT 4 TCGCrCGGTTT/ 3SpC3/ Adapter Adapter Primer
gtgactggagtt SEQ Primer cagacgtgtgct ID cttccgatctAA NO:
TGATACGGCGAC 5 CACCGAGATCTA CArCAAGGC/ 3SpC3/ P5 Adapter Example
Sequence AATGATACGGCG SEQ ACCACCGAGATC ID TACACTAGATCG NO:
CNNWNNWNNACA 6 CTCTTTCCCTAC ACGACGCTCTTC CGATC*T SP1 Sequencing
Primer 1 acactctttccc SEQ tacacgacgctc ID ttccgatct NO: 7 SP2
Sequencing Primer 2 gtgactggagtt SEQ cagacgtgtgct ID cttccgatct NO:
8 "*" indicates a phosphorothioate linkage; "rN" indicates a
ribonucleotide, where N is the nucleotide preceeded by the "r";
"/3SpC3/" indicates a 3'-C.sub.3 spacer.
[0033] One embodiment described herein is a method for identifying
and identifying and nominating on- and off-target CRISPR editing
sites with improved accuracy and sensitivity, the process
comprising the steps of: (a) co-delivering a guide sequence RNA
(sgRNA) or a two-part CRISPR RNA:trans-activating crRNA
(crRNA:tracrRNA) duplex and one or more tag sequences to cells; (b)
incubating the cells for a period of time; (c) isolating genomic
DNA from the cells, fragmenting the genomic DNA, and ligating the
fragmented genomic DNA to a unique molecular index containing a
universal adapter sequence; (d) amplifying the ligated DNA
fragments using primers targeting the tag and universal adapter
sequences to produce a first set of amplified sequences; (e)
amplifying the first set of amplified sequences using universal
sequencing primers targeting the tails of Tag-pTOP or Tag-pBOT
primers to produce a second set of amplified sequences; (f)
sequencing the pooled sequences and obtaining sequencing data; and
(g) identifying on-/off-target CRISPR editing loci. In one
embodiment, the universal sequencing primers target SP1 or SP2
sequence (SEQ ID NO: 7, 8) tails on the Tag-pTOP or Tag-pBOT
primers to produce a second set of amplified sequences. In another
embodiment, the universal sequencing primers target predesigned
non-homologous sequence (Table 6; SEQ ID NO: 269-273) tails on the
Tag-pTOP or Tag-pBot to produce a second set of amplified
sequences. In yet another embodiment, the universal primers target
predesigned 13-mer tails on the Tag-pTOP or Tag-pBOT primers to
produce a second set of amplified sequences. In one embodiment,
step (g) comprises executing on a processor: (i) aligning the
sequence data to a reference genome; (ii) identifying
on-/off-target CRISPR editing loci; and (iii) outputting the
alignment, analysis, and results data as tables or graphics. In
another embodiment, the method further comprises a step following
step (e) comprising: (e1) normalizing the second set of amplified
sequences to produce concentration normalized libraries, pooling
the normalized libraries with other samples to produce pooled
libraries; and continuing with steps (f)-(i). In one aspect, step
(d) uses a supression PCR method. In another aspect, the cells
constitutively express a Cas enzyme, are co-delivered with a Cas
expression vector, are co-delivered with a Cas protein, or are
co-delivered with a Cas RNP complex. In another aspect, the cells
constitutively express a Cas9 enzyme, are co-delivered with a Cas9
expression vector, are co-delivered with a Cas9 protein, or are
co-delivered with a Cas9 RNP complex. In another aspect, the cells
comprise human or mouse cells. In another aspect, the period of
time is about 24 hours to about 96 hours. In another aspect,
multiple tag sequences are co-delivered. In another aspect, the tag
sequences comprise double-stranded deoxyribooligonucleotides
(dsDNA) comprising 52-base pairs. In another aspect, the tag
sequences comprise a 5'-terminal phosphate, and phosphorothioate
linkages between the 1.sup.st and 2.sup.nd, 2.sup.nd and 3.sub.rd,
50.sup.th and 51.sup.st, and 51.sup.st and 52.sup.nd nucleotides.
In another aspect, the tag sequences comprise a double stranded DNA
comprising the top and bottom strand pairs of SEQ ID NO: 9-40 or
45-268.
[0034] Another embodiment described herein is on- and off-target
CRISPR editing sites identified or nominated using the methods
described herein.
[0035] Another embodiment described herein is a method for
designing 52-base pair tag sequences, the method comprising,
executing on a processor: (a) randomly generating 13-nucleotide
sequences with 40-90% GC content, max homopolymer length A:2, C:3,
G:2, T:2, weighted homopolymer rate <20, self-folding
T.sub.m<50.degree. C., and self-dimer T.sub.m<50.degree. C.;
(b) removing sequences that perfectly align to a particular genome
or that are homopolymers or GG or CC dinucleotide motifs and
obtaining a set of 13-mers; (c) selecting a subset of the 13-mer
sequences that contain one or less CC or GG dinucleotide motifs;
(d) concatenating four of the of 13-mer subset sequences to form
random 52-mer sequences; (e) aligning the random 52-mer sequences
to a genome; (f) removing the random 52-mer sequences that have
similarity to the genome to produce a subset of 52-mer sequences;
and (h) outputting the subset of 52-mer sequences and generating
the complementary strands to produce double stranded 52-base pair
tag sequences. In one aspect, the genome is human or mouse. In one
aspect, the 52-base pair tag sequences are not complementary to the
genome. In another aspect, the method further comprises designing
primers for the 52-base pair tag sequences. In another aspect, the
52-base pair tag sequences comprise a 5'-terminal phosphate, and
phosphorothioate linkages between the 1.sup.st and 2.sup.nd,
2.sup.nd and 3.sub.rd, 50.sup.th and 51.sup.st, and 51.sup.st and
52.sup.nd nucleotides of the 52-base pair tag sequences. In another
aspect, the method further comprises synthesising oligonucleotides
comprising the 52-base pair tag sequences, the complement of the
52-base pair tag sequences, or primers for the 52-base pair tag
sequences.
[0036] Another embodiment described herein is one or more 52-base
pair tag sequences designed using the methods described herein. In
one aspect, the 52-base pair tag sequence comprises a double
stranded DNA comprising the complementary top and bottom strand
pairs of SEQ ID NO: 9-40 or 45-268.
[0037] Another embodiment described herein is a method for
designing primers partially complementary to the 52-base pair tag
sequences described herein and an adapter primer, the method
comprising, executing on a processor: (a) designing tag primers
that are partially complementary to the top and bottom strands of
tag sequences; and (b) designing an adapter primer that is
partially complementary to the top strand of the adapter sequence;
wherein: the tag primers comprise a 5'-universal tail sequence
complementary to an SP1 or SP2 sequence (SEQ ID NO: 7, 8), a locus
specific segment, a ribonucleotide (rN) 6-nucleotides from the
3'-end, a 3'-end mismatch, and a 3'-end block (3'-C.sub.3 spacer);
and the adapter primer comprises a sequence complementary to the
SP1 or SP2 sequence (SEQ ID NO: 7, 8). In one aspect, the primers
partially complementary to top and bottom strands of the tag
sequences comprise a sequence complementary to the SP1 sequence and
the adapter primer comprises a sequence complementary to the SP2
sequence; or the primers partially complementary to top and bottom
strands of the tag sequences comprise a sequence complementary to
the SP2 sequence and the adapter primer comprises a sequence
complementary to the SP1 sequence. In another aspect, amplification
of a nucleic acid molecule with the primers that are complementary
to the top and bottom strands of tag sequences and primers that are
complementary to the top strand of the adapter sequence produces a
PCR product that comprises a portion of the tag sequence, a sgDNA
sequence, and the adapter sequence. In another aspect, the method
further comprises synthesising oligonucleotides comprising the
sequences of the forward and reverse tag primers and the adapter
primer.
[0038] In another embodiment described herein, the 52-base pair tag
sequences and primers partially complementary to the 52-base pair
tag sequences are designed and selected using an algorithm
predicting whether the primers are likely to be partially
complementary and have a propensity to form primer-dimers.
[0039] Another embodiment described herein is one or more primers
partially complementary to the 52-base pair tag sequences and one
or more adapter primers designed using the methods described
herein. In one aspect, the primers partially complementary to the
52-base pair tag sequence comprise the sequences of SEQ ID NO: 3,
4; and the adapter primer comprises the sequence of SEQ ID
NO:5.
[0040] Another embodiment described herein is the use of one or
more double-stranded 52-base pair tag sequences for identifying on-
and off-target CRISPR editing sites.
[0041] It will be apparent to one of ordinary skill in the relevant
art that suitable modifications and adaptations to the
compositions, formulations, methods, processes, and applications
described herein can be made without departing from the scope of
any embodiments or aspects thereof. The compositions and methods
provided are exemplary and are not intended to limit the scope of
any of the specified embodiments. All the various embodiments,
aspects, and options disclosed herein can be combined in any
variations or iterations. The scope of the methods and processes
described herein include all actual or potential combinations of
embodiments, aspects, options, examples, and preferences herein
described. The methods described herein may omit any component or
step, substitute any component or step disclosed herein, or include
any component or step disclosed elsewhere herein. It should also be
understood that embodiments may include and otherwise be
implemented by a combination of various hardware, software, and
electronic components. For example, various microprocessors and
application specific integrated circuits ("ASICs") can be utilized,
as can software of a variety of languages. Also, servers and
various computing devices can be used and can include one or more
processing units, one or more computer-readable mediums, one or
more input/output interfaces, and various connections (e.g., a
system bus) connecting the components. Should the meaning of any
terms in any of the patents or publications incorporated by
reference conflict with the meaning of the terms used in this
disclosure, the meanings of the terms or phrases in this disclosure
are controlling. Furthermore, the specification discloses and
describes merely exemplary embodiments. All patents and
publications cited herein are incorporated by reference herein for
the specific teachings thereof.
[0042] Various embodiments and aspects of the inventions described
herein are summarized by the following clauses: [0043] Clause 1. A
method for identifying and nominating on- and off-target CRISPR
edited sites with improved accuracy and sensitivity, the process
comprising the steps of: [0044] (a) co-delivering a guide sequence
RNA (sgRNA) or a two-part CRISPR RNA:trans-activating crRNA
(crRNA:tracrRNA) duplex, one or more tag sequences, and an
RNA-guided endonuclease to cells; [0045] (b) incubating the cells
for a period of time sufficient for double strand breaks to occur;
(c) isolating genomic DNA from the cells, fragmenting the genomic
DNA, and ligating the fragmented genomic DNA to a unique molecular
index containing a universal adapter sequence; [0046] (d)
amplifying the ligated DNA fragments using primers targeting the
tag and universal adapter sequences to produce a first set of
amplified sequences; [0047] (e) amplifying the first set of
amplified sequences using universal sequencing primers targeting
the tails of Tag-pTOP or Tag-pBOT primers to produce a second set
of amplified sequences; [0048] (f) sequencing the pooled sequences
and obtaining sequencing data; and [0049] (g) identifying
on-/off-target CRISPR editing loci. [0050] Clause 2. The method of
clause 1, wherein the universal sequencing primers target SP1 or
SP2 sequence (SEQ ID NO: 7, 8) tails on the Tag-pTOP or Tag-pBOT
primers to produce a second set of amplified sequences. [0051]
Clause 3. The method of clause 1 or 2, wherein the universal
sequencing primers target predesigned non-homologous sequence (SEQ
ID NO: 269-273) tails on the Tag-pTOP or Tag-pBot primers to
produce a second set of amplified sequences. [0052] Clause 4. The
method of any one of clauses 1-3, wherein the universal sequencing
primers target predesigned 13-mer tails on the Tag-pTOP or Tag-pBot
primers to produce a second set of amplified sequences. [0053]
Clause 5. The method of any one of clauses 1-4, wherein step (g)
comprises executing on a processor: [0054] Clause 6. aligning the
sequence data to a reference genome; [0055] (a) (ii) identifying
on-/off-target CRISPR editing loci; and [0056] (b) (iii) outputting
the alignment, analysis, and results data as custom-formatted
files, tables or graphics. [0057] Clause 7. The method of any one
of clauses 1-5, further comprising a step following step (e)
comprising: [0058] (a) (e1) normalizing the second set of amplified
sequences to produce concentration normalized libraries, pooling
the normalized libraries with other samples to produce pooled
libraries; and continuing with steps (f)-(i). [0059] Clause 8. The
method of any one of clauses 1-6, wherein step (d) uses a
supression PCR method. [0060] Clause 9. The method of any one of
clauses 1-7, wherein the RNA-guided endonuclease comprises an
endogenously-expressed Cas enzyme, a Cas expression vector, a Cas
protein, or a Cas RNP complex. [0061] Clause 10. The method of any
one of clauses 1-8, wherein the RNA-guided endonuclease comprises
an endogenously-expressed Cas9 enzyme, a Cas9 expression vector, a
Cas9 protein, or a Cas9 RNP complex. [0062] Clause 11. The method
of any one of clauses 1-9, wherein the cells comprise human or
mouse cells. [0063] Clause 12. The method of any one of clauses
1-10, wherein the period of time is about 24 hours to about 96
hours. [0064] Clause 13. The method of any one of clauses 1-11,
wherein multiple tag sequences are co-delivered. [0065] Clause 14.
The method of any one of clauses 1-12, wherein the tag sequences
comprise double-stranded deoxyribooligonucleotides (dsDNA)
comprising 52-base pairs. [0066] Clause 15. The method of any one
of clauses 1-13, wherein the tag sequences comprise a 5'-terminal
phosphate, and phosphorothioate linkages between the 1.sup.st and
2.sup.nd, 2.sup.nd and 3.sup.rd, 50.sup.th and 51.sup.st, and
51.sup.st and 52.sup.nd nucleotides. [0067] Clause 16. The method
of any one of clauses 1-14, wherein the tag sequences comprise a
double stranded DNA comprising the complementary top and bottom
strand pairs of SEQ ID NO: 1-2 or 7-268. [0068] Clause 17. On- and
off-target CRISPR editing sites identified or nominated using the
method of any one of clauses 1-15. [0069] Clause 18. A method for
designing 52-base pair tag sequences, the method comprising,
executing on a processor: [0070] (a) randomly generating
13-nucleotide sequences with 40-90% GC content, max homopolymer
length A:2, C:3, G:2, T:2, weighted homopolymer rate <20,
self-folding T.sub.m<50.degree. C., and self-dimer
T.sub.m<50.degree. C.; [0071] (b) removing sequences that
perfectly align to a particular genome or that are homopolymers or
GG or CC dinucleotide motifs and obtaining a set of 13-mers; [0072]
(c) selecting a subset of the 13-mer sequences that contain one or
less CC or GG dinucleotide motifs; [0073] (d) concatenating four of
the of 13-mer subset sequences to form random 52-mer sequences;
[0074] (e) aligning the random 52-mer sequences to a genome; [0075]
(f) removing the random 52-mer sequences that have similarity to
the genome to produce a subset of 52-mer sequences; and [0076] (g)
outputting the subset of 52-mer sequences and generating the
complementary strands to produce double stranded 52-base pair tag
sequences. [0077] Clause 19. The method of clause 17, wherein the
genome is human or mouse. [0078] Clause 20. The method of clause 17
or 18, wherein the 52-base pair tag sequences are-non complementary
to the genome. [0079] Clause 21. The method of any one of clauses
17-19, further comprising designing primers for the 52-base pair
tag sequences. [0080] Clause 22. The method of any one of clauses
17-20, wherein the 52-base pair tag sequences comprise a
5'-terminal phosphate, and phosphorothioate linkages between the
1.sup.st and 2.sup.nd, 2.sup.nd and 3.sup.rd, 50.sup.th and
51.sup.st, and 51.sup.st and 52.sup.nd nucleotides of the 52-base
pair tag sequences. [0081] Clause 23. The method of any one of
clauses 17-21, further comprising synthesizing oligonucleotides
comprising the 52-base pair tag sequences, the complement of the
52-base pair tag sequences, or primers for the 52-base pair tag
sequences. [0082] Clause 24. One or more 52-base pair tag sequences
designed using the methods of clauses 17-22. [0083] Clause 25. The
52-base pair tag sequences of clause 23, wherein the 52-base pair
tag sequence comprises a double stranded DNA comprising the top and
bottom strand pairs of SEQ ID NO: 1-2 or 7-268. [0084] Clause 26. A
method for designing primers partially complementary to the 52-base
pair tag sequences of clause 23 and an adapter primer, the method
comprising, executing on a processor: [0085] (a) designing tag
primers that are partially complementary to the top and bottom
strands of tag sequences; and [0086] (b) designing an adapter
primer that is partially complementary to the top strand of the
adapter sequence; [0087] (c) wherein: [0088] (d) the tag primers
comprise a 5'-universal tail sequence; and [0089] (e) the adapter
primer comprises a sequence complementary to the tails of Tag-pTOP
or Tag-pBOT primers. [0090] Clause 27. The method of clause 25,
wherein the 5'-universal tail sequence is complementary to an SP1
or SP2 sequence (SEQ ID NO: 7, 8), a locus specific segment, a
ribonucleotide (rN) 6-nucleotides from the 3'-end, a 3'-end
mismatch, a 3'-end block (3'-C.sub.3 spacer), a predesigned
non-homologous sequence (SEQ ID NO: 269-273), or a predesigned
13-mer sequence. [0091] Clause 28. The method of clause 25 or 26,
wherein the primers partially complementary to top and bottom
strands of the tag sequences comprise a tail sequence complementary
to the SP1 sequence (SEQ ID NO: 7) and the adapter primer comprises
a sequence complementary to the SP2 sequence (SEQ ID NO: 8) tail on
the Tag-pTOP or Tag-pBOT primers; or the primers partially
complementary to top and bottom strands of the tag sequences
comprise a tail sequence complementary to the SP2 sequence (SEQ ID
NO: 8) and the adapter primer comprises a sequence complementary to
the SP1 sequence (SEQ ID NO: 7) tail on the Tag-pTOP or Tag-pBOT
primers. [0092] Clause 29. The method of any one of clauses 25-27,
wherein the amplification of a nucleic acid molecule with the
primers that are complementary to the top and bottom strands of tag
sequences and primers that are complementary to the top strand of
the adapter sequence produces a PCR product that comprises a
portion of the tag sequence, a sgDNA sequence, and the adapter
sequence. [0093] Clause 30. The method of any one of clauses 25-28,
further comprising synthesizing oligonucleotides comprising the
sequences of the forward and reverse tag primers and the adapter
primer. [0094] Clause 31. The method of any one of clauses 17-21
and 25-29, wherein the 52-base pair tag sequences and primers
partially complementary to the 52-base pair tag sequences are
designed and selected using an algorithm predicting whether the
primers are likely to be partially complementary and have a
propensity to form primer-dimers. [0095] Clause 32. One or more
primers partially complementary to the 52-base pair tag sequences
and one or more adapter primers designed using the method of
clauses 22-25. [0096] Clause 33. The primers of clause 32, wherein
the primers comprise the sequences of SEQ ID NO: 3, 4; and the
adapter primer, wherein the adapter primer comprises the sequence
of SEQ ID NO: 5. [0097] Clause 34. Use of one or more
double-stranded 52-base pair tag sequences for identifying on- and
off-target CRISPR editing sites.
REFERENCES
[0097] [0098] 1. Wenert et al., "Unbiased detection of CRISPR
off-targets in vivo using DISCOVER-seq," Science 364(6437): 286-289
(2019). [0099] 2. Nobles et al., "IGUIDE: An improved pipeline for
analyzing CRISPR cleavage specificity," Genome Biol. 20(14): 4-9
(2019). [0100] 3. Tsai et al., "GUIDE-seq enables genome-wide
profiling of off-target cleavage by CRISPR-Cas nucleases," Nature
Biotechnol. 33(2): 187-197 (2015). [0101] 4. Yan et al., "BLISS is
a versatile and quantitative method for genome-wide profiling of
DNA double-strand breaks," Nature Commun. 8: 15058 (2017). [0102]
5. Tsai et al., "CIRCLE-seq: a highly sensitive in vitro screen for
genome-wide CRISPR-Cas9 nuclease off-targets," Nature Methods
14(6): 607-614 (2017). [0103] 6. Cameron et al., "Mapping the
genomic landscape of CRISPR-Cas9 cleavage," Nature Methods 14(6):
600-606 (2017). [0104] 7 Char and Moosburner, "Unraveling
CRISPR-Cas9 genome engineering parameters via a library-on-library
approach," Nature Methods 12(9): 823-826 (2015). [0105] 8. Rand et
al., "Headloop suppression PCR and its application to selective
amplification of methylated DNA sequences," Nucleic Acids Res.
33(14):e127 (2005).
EXAMPLES
Example 1
[0106] This experiment demonstrates the increased efficiency in tag
integration when using double-stranded DNA tags with a length of
52-base pairs and varying genetic sequence. The sequences used are
shown in Tables 3-5. Double-stranded tags were generated by
hybridization of a top strand and a complementary bottom strand
(Tables 3-4; SEQ ID NO: 9-40 or 45-268). Sixteen different tag
designs were introduced separately into HEK293 cells constitutively
expressing Cas9 together with a guideRNA which targets the EMX1
locus. Alternatively, either pools of 16 tags or one pool of 112
tags were introduced into HEK293 cells constitutively expressing
Cas9 together with a guideRNA which targets the EMX1 locus.
GuideRNAs were electroporated at a concentration of 10 .mu.M,
whereas the single Tag or pooled Tags were delivered at a final
concentration of 0.5 .mu.M. Tag integration levels were determined
by targeted amplification using rhAmpSeq primers (SEQ ID NO: 3-4),
enriching for known on- and off-target sites of the EMX1 guideRNA.
The rhAmpSeq pool for EMX1 consists of 32 sites, which represent
empirically determined ON and OFF target loci. Amplified products
were sequenced on an Illumina.RTM. MiSeq, and tag integration
levels were determined using custom software. This example shows
that tag integration efficiency varies among single tag constructs
individually with a range between 6 (CTL021) and 13 (CTL169,
CTL079, CTL002) sites out of a maximum of 32 sites, and is
therefore sequence dependent (Single Tags, FIG. 7). By taking the
mathematical union of the single tag results, a hypothetical number
of 23 sites was calculated (CTLmax, FIG. 7). The hypothesis that
combining a pool of tags would increase the likelihood of tag
integration was tested and was demonstrated (Pooled Tags, Table,
FIG. 7). Pool A1 consists of the tags represented in the Single
Tags (see Table 5) and demonstrated that 21 tag integration events
were detected out of a maximum of 32 sites, which is higher than
achieved with any of the single tags. Similarly, Pool B3
demonstrated integration of a tag at 21 sites out of a maximum of
32 sites. Again, variability between pools was shown (Pooled Tags,
FIG. 7), indicating optimization of tag designs can potentially
maximize tag integration.
TABLE-US-00003 TABLE 3 Sequences Used for Second Proof of Concept
SEQ ID Name Sequence (5'.fwdarw.3') NO CTL085_
/5Phos/A*C*GAGCGGTAGTCACCTA SEQ TOP_tag GTCGTCGTACCAATTCGACGCACACTA
ID CTCGC*G*C NO: 9 CTL085_ /5Phos/G*C*GCGAGTAGTGTGCGTC SEQ BOT_tag
GAATTGGTACGACGACTAGGTGACTAC ID CGCTC*G*T NO: 10 CTL169_
/5Phos/T*A*GCGCGAGTAGTCGGAC SEQ TOP_tag GAGCGGTTACCAATACGCCGCACCTTA
ID ATCCG*C*G NO: 11 CTL169_ /5Phos/C*G*CGGATTAAGGTGCGGC SEQ BOT_tag
GTATTGGTAACCGCTCGTCCGACTACT ID CGCGC*T*A NO: 12 CTL137_
/5Phos/T*C*GCGACAGTAGTCGTTC SEQ TOP_tag GGCTAGGTACCTATTACCGCGTAGTTA
ID GCGGC*G*T NO: 13 CTL137_ /5Phos/A*C*GCCGCTAACTACGCGG SEQ BOT_tag
TAATAGGTACCTAGCCGAACGACTACT ID GTCGC*G*A NO: 14 CTL042_
/5Phos/C*G*CGCTACTAGGTGCGTC SEQ TOP_tag GAATTGGTACCGATCCGCAATACACTA
ID CTCGC*G*C NO: 15 CTL042_ /5Phos/G*C*GCGAGTAGTGTATTGC SEQ BOT_tag
GGATCGGTACCAATTCGACGCACCTAG ID TAGCG*C*G NO: 16 CTL051_
/5Phos/G*G*TAACGAGCGGTGCGTC SEQ TOP_tag GAATTGGTAACCGCTCGTCCGACCTTA
ID ATCGC*G*C NO: 17 CTL051_ /5Phos/G*C*GCGATTAAGGTCGGAC SEQ BOT_tag
GAGCGGTTACCAATTCGACGCACCGCT ID CGTTA*C*C NO: 18 CTL167_
/5Phos/T*T*CGGCGCTAGGTGCGGC SEQ TOP_tag GTATTGGTAACCGCTCGTCCGTTCGGC
ID GCTAG*G*T NO: 19 CTL167_ /5Phos/A*C*CTAGCGCCGAACGGAC SEQ BOT_tag
GAGCGGTTACCAATACGCCGCACCTAG ID CGCCG*A*A NO: 20 CTL026_
/5Phos/T*A*CGCGACTAGGTGCGCG SEQ TOP_tag ATTAAGGTACCTATTACCGCGCGACTA
ID TGTGC*G*C NO: 21 CTL026_ /5Phos/G*C*GCACATAGTCGCGCGG SEQ BOT_tag
TAATAGGTACCTTAATCGCGCACCTAG ID TCGCG*T*A NO: 22 CTL068_
/5Phos/G*T*CGCGCAGTGTAGCGCG SEQ TOP_tag ATTAAGGTACCTATTACCGCGTCGCGA
ID CAGTA*G*T NO: 23 CTL068_ /5Phos/A*C*TACTGTCGCGACGCGG SEQ BOT_tag
TAATAGGTACCTTAATCGCGCTACACT ID GCGCG*A*C NO: 24 CTL138_
/5Phos/A*A*CCGTCGATCCGCGCGT SEQ TOP_tag AGTATGGTACCGATCCGCAATACTAGC
ID GCGAC*A*A NO: 25 CTL138_ /5Phos/T*T*GTCGCGCTAGTATTGC SEQ BOT_tag
GGATCGGTACCATACTACGCGCGGATC ID GACGG*T*T NO: 26 CTL079_
/5Phos/T*C*GCTCGATTGGTTACGC SEQ TOP_tag GCACTACTTATGCGCTCGACTCGTTCG
ID GCTAG*G*T NO: 27 CTL079_ /5Phos/A*C*CTAGCCGAACGAGTCG SEQ BOT_tag
AGCGCATAAGTAGTGCGCGTAACCAAT ID CGAGC*G*A NO: 28 CTL063_
/5Phos/A*C*TGCGAGCGTACTTGTC SEQ TOP_tag GCGCTAGTACCAATTCGACGCAACCGC
ID TCGTC*C*G NO: 29 CTL063_ /5Phos/C*G*GACGAGCGGTTGCGTC SEQ BOT_tag
GAATTGGTACTAGCGCGACAAGTACGC ID TCGCA*G*T NO: 30 CTL168_
/5Phos/C*G*CATTAGTCGGTGCGGC SEQ TOP_tag GTATTGGTAACCGCTCGTCCGACGCGC
ID TACCT*A*T NO: 31 CTL168_ /5Phos/A*T*AGGTAGCGCGTCGGAC SEQ BOT_tag
GAGCGGTTACCAATACGCCGCACCGAC ID TAATG*C*G NO: 32 CTL021_
/5Phos/A*T*TGCGGATCGGTGCGTC SEQ TOP_tag GAATTGGTAACCGCTCGTCCGTACGCG
ID CACTA*C*T NO: 33 CTL021_ /5Phos/A*G*TAGTGCGCGTACGGAC SEQ BOT_tag
GAAGCGGTTACCAATTCGCGCACCGAT ID CCGCA*A*T NO: 34 CTL151_
/5Phos/T*C*GGCGAGTAGTTGCGCG SEQ TOP_tag GTTATGGTACCATAACCGCGCAGTAGT
ID ACGCG*G*T NO: 35 CTL151_ /5Phos/A*C*CGCGTACTACTGCGCG SEQ BOT_tag
GTTATGGTACCATAACCGCGCAACTAC ID TCGCC*G*A NO: 36 CTL002_
/5Phos/A*C*TAGCGATCGGTACCTA SEQ TOP_tag GCGCCGAAACCTATTACCGCGACCTAG
ID CGTTG*C*G NO: 37 CTL002_ /5Phos/C*G*CAACGCTAGGTCGCGG SEQ BOT_tag
TAATAGGTTTCGGCGCTAGGTACCGAT ID CGCTA*G*T NO: 38 CTL134_
/5Phos/T*A*GCGCGTCAAGAGCGCG SEQ TOP_tag GTTATGGTTTCGGCGCTAGGTTAACAG
ID CGCGT*C*G NO: 39 CTL134_ /5Phos/C*G*ACGCGCTGTTAACCTA SEQ BOT_tag
GCGCCGAAACCATAACCGCGCTCTTGA ID CGCGC*T*A NO: 40 GuideSeq_
/5Phos/G*T*TTAATTGAGTTGTCAT SEQ TOP_tag ATGTTAATAACGGT*A*T ID NO:
41 GuideSeq_ /5Phos/A*T*ACCGTTATTAACATAT SEQ BOT_tag
GACAACTCAATTAA*A*C ID NO: 42 EMX1 GAGTCCGAGCAGAAGAAGAA SEQ
protospacer ID NO: 43 AR GTTGGAGCATCTGAGTCCAG SEQ protospacer ID
NO: 44 "/5Phos/" indicates a 5'-phosphate moiety; "*" indicates a
phosphorothioate linkage.
Example 2
[0107] This experiment demonstrates the increased efficiency in tag
integration when using double-stranded DNA tags with a length of
52-base pairs and varying genetic sequence. The sequences used are
shown in Tables 3-5. Double-stranded tags were generated by
hybridization of a top strand and a complementary bottom strand
(SEQ ID NO: 9-40 or 45-268). Sixteen different tag designs were
introduced separately into HEK293 cells constitutively expressing
Cas9 together with a guideRNA which targets the AR locus.
Alternatively, either pools of 16 tags or one pool of 112 tags were
introduced into HEK293 cells constitutively expressing Cas9
together with a guideRNA which targets the AR locus. GuideRNAs were
electroporated at a concentration of 10 .mu.M, whereas the single
Tag or pooled Tags were delivered at a final concentration of 0.5
.mu.M. Tag integration levels were determined by targeted
amplification using rhAmpSeq primers (SEQ ID NO: 3-4), enriching
for known on- and off-target sites of the AR guideRNA. The rhAmpSeq
pool for AR consists of 53 sites which represent empirically
determined ON and OFF target loci. Amplified products were
sequenced on an Illumina.RTM. MiSeq, and tag integration levels
were determined using custom software. This example shows that tag
integration efficiency varies among single tag constructs
individually with a range between 35 (CTL085, CTL134) and 41 sites
(CTL002) out of a maximum of 53 sites, and is therefore sequence
dependent (Single Tags, Table 5, FIG. 8).
[0108] By taking the mathematical union of the single tag results,
a hypothetical number of 47 sites was calculated (CTLmax, FIG. 8).
The hypothesis that combining a pool of tags would increase the
likelihood of tag integration was tested and was demonstrated
(Pooled Tags, Table 5, FIG. 8). Pool B4 (see Table 5) demonstrated
that 44 tag integration events were detected out of a maximum of 53
sites, which is higher than achieved with any of the single tags.
Again, variability between pools was shown (Pooled Tags, Table 5,
FIG. 8), indicating optimization of tag designs can potentially
maximize tag integration.
TABLE-US-00004 TABLE 4 Tag Sequences Name Sequence (5'.fwdarw.3')
SEQ ID NO CTL085_TOP_tag
/5Phos/A*C*GAGCGGTAGTCACCTAGTCGTCGTACCAATTCGA SEQ ID NO: 45
CGCACACTACTCGC*G*C CTL169_TOP_tag
/5Phos/T*A*GCGCGAGTAGTCGGACGAGCGGTTACCAATACGC SEQ ID NO: 46
CGCACCTTAATCCG*C*G CTL137_TOP_tag
/5Phos/T*C*GCGACAGTAGTCGTTCGGCTAGGTACCTATTACC SEQ ID NO: 47
GCGTAGTTAGCGGC*G*T CTL042_TOP_tag
/5Phos/C*G*CGCTACTAGGTGCGTCGAATTGGTACCGATCCGC SEQ ID NO: 48
AATACACTACTCGC*G*C CTL051_TOP_tag
/5Phos/G*G*TAACGAGCGGTGCGTCGAATTGGTAACCGCTCGT SEQ ID NO: 49
CCGACCTTAATCGC*G*C CTL167_TOP_tag
/5Phos/T*T*CGGCGCTAGGTGCGGCGTATTGGTAACCGCTCGT SEQ ID NO: 50
CCGTTCGGCGCTAG*G*T CTL026_TOP_tag
/5Phos/T*A*CGCGACTAGGTGCGCGATTAAGGTACCTATTACC SEQ ID NO: 51
GCGCGACTATGTGC*G*C CTL068_TOP_tag
/5Phos/G*T*CGCGCAGTGTAGCGCGATTAAGGTACCTATTACC SEQ ID NO: 52
GCGTCGCGACAGTA*G*T CTL138_TOP_tag
/5Phos/A*A*CCGTCGATCCGCGCGTAGTATGGTACCGATCCGC SEQ ID NO: 53
AATACTAGCGCGAC*A*A CTL079_TOP_tag
/5Phos/T*C*GCTCGATTGGTTACGCGCACTACTTATGCGCTCG SEQ ID NO: 54
ACTCGTTCGGCTAG*G*T CTL063_TOP_tag
/5Phos/A*C*TGCGAGCGTACTTGTCGCGCTAGTACCAATTCGA SEQ ID NO: 55
CGCAACCGCTCGTC*C*G CTL168_TOP_tag
/5Phos/C*G*CATTAGTCGGTGCGGCGTATTGGTAACCGCTCGT SEQ ID NO: 56
CCGACGCGCTACCT*A*T CTL021_TOP_tag
/5Phos/A*T*TGCGGATCGGTGCGTCGAATTGGTAACCGCTCGT SEQ ID NO: 57
CCGTACGCGCACTA*C*T CTL151_TOP_tag
/5Phos/T*C*GGCGAGTAGTTGCGCGGTTATGGTACCATAACCG SEQ ID NO: 58
CGCAGTAGTACGCG*G*T CTL002_TOP_tag
/5Phos/A*C*TAGCGATCGGTACCTAGCGCCGAAACCTATTACC SEQ ID NO: 59
GCGACCTAGCGTTG*C*G CTL134_TOP_tag
/5Phos/T*A*GCGCGTCAAGAGCGCGGTTATGGTTTCGGCGCTA SEQ ID NO: 60
GGTTAACAGCGCGT*C*G CTL085_BOT_tag
/5Phos/G*C*GCGAGTAGTGTGCGTCGAATTGGTACGACGACTA SEQ ID NO: 61
GGTGACTACCGCTC*G*T CTL169_BOT_tag
/5Phos/C*G*CGGATTAAGGTGCGGCGTATTGGTAACCGCTCGT SEQ ID NO: 62
CCGACTACTCGCGC*T*A CTL137_BOT_tag
/5Phos/A*C*GCCGCTAACTACGCGGTAATAGGTACCTAGCCGA SEQ ID NO: 63
ACGACTACTGTCGC*G*A CTL042_BOT_tag
/5Phos/G*C*GCGAGTAGTGTATTGCGGATCGGTACCAATTCGA SEQ ID NO: 64
CGCACCTAGTAGCG*C*G CTL051_BOT_tag
/5Phos/G*C*GCGATTAAGGTCGGACGAGCGGTTACCAATTCGA SEQ ID NO: 65
CGCACCGCTCGTTA*C*C CTL167_BOT_tag
/5Phos/A*C*CTAGCGCCGAACGGACGAGCGGTTACCAATACGC SEQ ID NO: 66
CGCACCTAGCGCCG*A*A CTL026_BOT_tag
/5Phos/G*C*GCACATAGTCGCGCGGTAATAGGTACCTTAATCG SEQ ID NO: 67
CGCACCTAGTCGCG*T*A CTL068_BOT_tag
/5Phos/A*C*TACTGTCGCGACGCGGTAATAGGTACCTTAATCG SEQ ID NO: 68
CGCTACACTGCGCG*A*C CTL138_BOT_tag
/5Phos/T*T*GTCGCGCTAGTATTGCGGATCGGTACCATACTAC SEQ ID NO: 69
GCGCGGATCGACGG*T*T CTL079_BOT_tag
/5Phos/A*C*CTAGCCGAACGAGTCGAGCGCATAAGTAGTGCGC SEQ ID NO: 70
GTAACCAATCGAGC*G*A CTL063_BOT_tag
/5Phos/C*G*GACGAGCGGTTGCGTCGAATTGGTACTAGCGCGA SEQ ID NO: 71
CAAGTACGCTCGCA*G*T CTL168_BOT_tag
/5Phos/A*T*AGGTAGCGCGTCGGACGAGCGGTTACCAATACGC SEQ ID NO: 72
CGCACCGACTAATG*C*G CTL021_BOT_tag
/5Phos/A*G*TAGTGCGCGTACGGACGAGCGGTTACCAATTCGA SEQ ID NO: 73
CGCACCGATCCGCA*A*T CTL151_BOT_tag
/5Phos/A*C*CGCGTACTACTGCGCGGTTATGGTACCATAACCG SEQ ID NO: 74
CGCAACTACTCGCC*G*A CTL002_BOT_tag
/5Phos/C*G*CAACGCTAGGTCGCGGTAATAGGTTTCGGCGCTA SEQ ID NO: 75
GGTACCGATCGCTA*G*T CTL134_BOT_tag
/5Phos/C*G*ACGCGCTGTTAACCTAGCGCCGAAACCATAACCG SEQ ID NO: 76
CGCTCTTGACGCGC*T*A CTL161_TOP_tag
/5Phos/T*A*CACTGCGCGACACTGCGAGCGTACACCTTAATCG SEQ ID NO: 77
CGCTAGTTAGCGGC*G*T CTL164_TOP_tag
/5Phos/A*A*CCGTCGAGTGCACCGCGTACTACTAATGTCGAAC SEQ ID NO: 78
CGCTACGCGCACTA*C*T CTL030_TOP_tag
/5Phos/C*G*CGGACTAAGGTGCGCGAGTAGTGTTACGCGCACT SEQ ID NO: 79
ACTAATCTAGCCGC*G*A CTL088_TOP_tag
/5Phos/A*C*TAGTGCGACGAACTACTCGCGCTAACCAATTCGA SEQ ID NO: 80
CGCACCGATCGCTA*G*T CTL148_TOP_tag
/5Phos/A*A*TGTCGAACCGCGCGCGAGTAGTGTACCATAACCG SEQ ID NO: 81
CGCACCTTAGTCCG*C*G CTL152_TOP_tag
/5Phos/G*C*GTCGAATTGGTACCGCCGACTTATACCAATACGC SEQ ID NO: 82
CGCATAGGTAGCGC*G*T CTL007_TOP_tag
/5Phos/A*C*CTAGTAGCGCGGCGTCGAATTGGTACTAGCGCGA SEQ ID NO: 83
CAACGCGTAGTATG*G*T CTL141_TOP_tag
/5Phos/A*C*CGCTCGTTACCGCGCGATTAAGGTACGCCGCTAA SEQ ID NO: 84
CTACGGTACGGTCG*G*T CTL064_TOP_tag
/5Phos/A*C*CGCCGACTTATCGTTCGGCTAGGTACCAATTCGA SEQ ID NO: 85
CGCACTGCGAGCGT*A*C CTL158_TOP_tag
/5Phos/A*C*CTTAATCCGCGACTGCGAGCGTACACCTATTACC SEQ ID NO: 86
GCGCGACGCGCTGT*T*A CTL066_TOP_tag
/5Phos/A*C*GACGACTAGGTACCGCTCGTTACCTCTTGACGCG SEQ ID NO: 87
CTAACCAATTCGAC*G*C CTL144_TOP_tag
/5Phos/A*C*CATACTACGCGGCGGTTCGACATTACCATAACCG SEQ ID NO: 88
CGCTAGTGCGAGCG*T*A CTL107_TOP_tag
/5Phos/C*T*TGTACGGCGGTGCGGCGTATTGGTACCAATACGC SEQ ID NO: 89
CGCTCGTCGCACTA*G*T CTL149_TOP_tag
/5Phos/G*T*ACGCTCGCAGTACCGCCGACTTATACCTTAATCG SEQ ID NO: 90
CGCACTAGCGCGAC*A*A CTL008_TOP_tag
/5Phos/A*C*GACGACTAGGTTATGGTACGGCGTTAGCGCGAGT SEQ ID NO: 91
AGTACCTTAGTCCG*C*G CTL099_TOP_tag
/5Phos/A*C*GAGCGGTAGTCATAGGTAGCGCGTTCTTGACGCG SEQ ID NO: 92
CTAACCGATCGCTA*G*T CTL089_TOP_tag
/5Phos/A*C*CGATCCGCAATGCGTCGAATTGGTACCATAACCG SEQ ID NO: 93
CGCACCGCCGTACA*A*G CTL081_TOP_tag
/5Phos/A*C*TAGTGCGACGAACTACTGTCGCGAACCTATTACC SEQ ID NO: 94
GCGACCAATCGAGC*G*A CTL075_TOP_tag
/5Phos/A*C*CGCCGTACAAGTCGCGACAGTAGTAACCGCTCGT SEQ ID NO: 95
CCGTTCGGCGCTAG*G*T CTL160_TOP_tag
/5Phos/T*C*GTCGCACTAGTCGCATTAGTCGGTAGTAGTACGC SEQ ID NO: 96
GGTATAGGTAGCGC*G*T CTL133_TOP_tag
/5Phos/A*C*CAATTCGACGCTAGTTAGCGGCGTACACTACTCG SEQ ID NO: 97
CGCGCACTCGACGG*T*T CTL076_TOP_tag
/5Phos/C*G*CGGTAATAGGTCGCGGTAATAGGTACGAGCGGTA SEQ ID NO: 98
GTCACACTACTCGC*G*C CTL024_TOP_tag
/5Phos/T*C*GGCGAGTAGTTTAGTGCGAGCGTAAGTAGTGCGC SEQ ID NO: 99
GTAACCAATCGAGC*G*A CTL045_TOP_tag
/5Phos/G*T*CGCGCAGTGTAGCGCGGTTATGGTACCATAACCG SEQ ID NO: 100
CGCACTAGTGCGAC*G*A CTL009_TOP_tag
/5Phos/T*A*TGCGCTCGACTGCGCGATTAAGGTAATGTCGAAC SEQ ID NO: 101
CGCAGTAGTACGCG*G*T CTL055_TOP_tag
/5Phos/A*C*TAGCGCGACAACGACTATGTGCGCACCAATTCGA SEQ ID NO: 102
CGCTACGCGCACTA*C*T CTL101_TOP_tag
/5Phos/A*A*CTACTCGCCGACTTGTACGGCGGTACCAATTCGA SEQ ID NO: 103
CGCAACTAATCCGC*G*C CTL135_TOP_tag
/5Phos/C*G*CGGATTAAGGTCTTGTACGGCGGTACCTAGCCGA SEQ ID NO: 104
ACGTACGCGCACTA*C*T CTL155_TOP_tag
/5Phos/T*A*GCGCGTCAAGACTTGTACGGCGGTACCGATCCGC SEQ ID NO: 105
AATGCACTCGACGG*T*T CTL122_TOP_tag
/5Phos/C*G*CATTAGTCGGTGCGGCGTATTGGTACGACGACTA SEQ ID NO:
106 GGTACCAATACGCC*G*C CTL080_TOP_tag
/5Phos/A*C*CTAGTAGCGCGGCGCGGTTATGGTACCGACTAAT SEQ ID NO: 107
GCGACTAGCGATCG*G*T CTL126_TOP_tag
/5Phos/A*C*TACTCGCGCTAACCTAGTCGTCGTAATCTAGCCG SEQ ID NO: 108
CGATACGCTCGCAC*T*A CTL098_TOP_tag
/5Phos/A*C*CGCCGCTATACGCGCGATTAAGGTGTACGCTCGC SEQ ID NO: 109
AGTCGCGGACTAAG*G*T CTL038_TOP_tag
/5Phos/T*A*CGCGCACTACTAACCGTCGAGTGCGTACGCTCGC SEQ ID NO: 110
AGTACCGATCGCTA*G*T CTL139_TOP_tag
/5Phos/G*T*CGCGCAGTGTATAACAGCGCGTCGTTAGTGCGCG SEQ ID NO: 111
AGAACGACGACTAG*G*T CTL010_TOP_tag
/5Phos/G*C*GTCGAATTGGTCGCGTAGTATGGTACCGCCGCTA SEQ ID NO: 112
TACACCAATACGCC*G*C CTL034_TOP_tag
/5Phos/T*A*CGCGCACTACTTACGCGACTAGGTACCGATCGCT SEQ ID NO: 113
AGTCGACGCGCTGT*T*A CTL117_TOP_tag
/5Phos/A*C*GCCGCTAACTATAGTTAGCGGCGTACCAATTCGA SEQ ID NO: 114
CGCAACTAATCCGC*G*C CTL035_TOP_tag
/5Phos/C*G*CGGACTAAGGTTAGTTAGCGGCGTTACGCGCACT SEQ ID NO: 115
ACTACCGATCCGCA*A*T CTL121_TOP_tag
/5Phos/A*C*GACGACTAGGTACCGCCGACTTATACGCCGCTAA SEQ ID NO: 116
CTAATAGGTAGCGC*G*T CTL106_TOP_tag
/5Phos/C*G*GATCGACGGTTGCGCGAGTAGTGTAGTAGTACGC SEQ ID NO: 117
GGTTACACTGCGCG*A*C CTL059_TOP_tag
/5Phos/A*T*TGCGGATCGGTACCGCCGACTTATACCGATCCGC SEQ ID NO: 118
AATTCGCTCGATTG*G*T CTL157_TOP_tag
/5Phos/A*C*TGCGAGCGTACACTGCGAGCGTACACCTTAATCG SEQ ID NO: 119
CGCACCGCTCGTTA*C*C CTL015_TOP_tag
/5Phos/A*C*TACTGTCGCGATCGTCGCACTAGTTACGCTCGCA SEQ ID NO: 120
CTAATTGCGGATCG*G*T CTL110_TOP_tag
/5Phos/G*G*TAACGAGCGGTTCTCGCGCACTAATTAGTGCGCG SEQ ID NO: 121
AGAACCATACTACG*C*G CTL123_TOP_tag
/5Phos/A*C*TACTCGCGCTAGCGCGATTAAGGTACCTTAATCG SEQ ID NO: 122
CGCAACTACTCGCC*G*A CTL014_TOP_tag
/5Phos/T*A*CGCGCACTACTCTTGTACGGCGGTACCAATTCGA SEQ ID NO: 123
CGCAACCGTCGAGT*G*C CTL131_TOP_tag
/5Phos/A*A*CCGTCGATCCGATTGCGGATCGGTACCTTAATCG SEQ ID NO: 124
CGCACTAGTGCGAC*G*A CTL062_TOP_tag
/5Phos/A*G*TAGTGCGCGTATACACTGCGCGACACACTACTCG SEQ ID NO: 125
CGCACCTTAATCCG*C*G CTL044_TOP_tag
/5Phos/A*C*GCCGTACCATACGCGGTAATAGGTAGTAGTGCGC SEQ ID NO: 126
GTATTCGGCGCTAG*G*T CTL043_TOP_tag
/5Phos/T*A*GCGCGTCAAGAACCTAGCGTTGCGATAAGTCGGC SEQ ID NO: 127
GGTAGTAGTACGCG*G*T CTL118_TOP_tag
/5Phos/C*G*CATTAGTCGGTAATCTAGCCGCGAACCATAACCG SEQ ID NO: 128
CGCACCGATCGCTA*G*T CTL128_TOP_tag
/5Phos/T*A*TGGTACGGCGTGCGGCGTATTGGTACGCCGCTAA SEQ ID NO: 129
CTAATAAGTCGGCG*G*T CTL067_TOP_tag
/5Phos/G*C*GCGGTTATGGTGCGGCGTATTGGTACGAGCGGTA SEQ ID NO: 130
GTCAACCGCTCGTC*C*G CTL020_TOP_tag
/5Phos/C*G*ACTATGTGCGCAACTACTCGCCGAACCATAACCG SEQ ID NO: 131
CGCTATGCGCTCGA*C*T CTL006_TOP_tag
/5Phos/T*A*GTTAGCGGCGTACCGCTCGTTACCACCTTAATCG SEQ ID NO: 132
CGCACCATACTACG*C*G CTL017_TOP_tag
/5Phos/C*G*CATTAGTCGGTAGTAGTGCGCGTAAACCGCTCGT SEQ ID NO: 133
CCGTTAGTGCGCGA*G*A CTL057_TOP_tag
/5Phos/T*A*GCGCGAGTAGTACCGACTAATGCGTCTCGCGCAC SEQ ID NO: 134
TAAGACTACCGCTC*G*T CTL078_TOP_tag
/5Phos/T*A*CGCTCGCACTATCGCTCGATTGGTACCGCCGCTA SEQ ID NO: 135
TACACCATAACCGC*G*C CTL031_TOP_tag
/5Phos/A*C*CAATCGAGCGAAGTCGAGCGCATAACGCGCTACC SEQ ID NO: 136
TATACGCCGCTAAC*T*A CTL136_TOP_tag
/5Phos/A*C*CTTAATCCGCGACTGCGAGCGTACACCGACTAAT SEQ ID NO: 137
GCGACTACTGTCGC*G*A CTL165_TOP_tag
/5Phos/A*G*TAGTGCGCGTATCGCTCGATTGGTTCTTGACGCG SEQ ID NO: 138
CTAGTATAGCGGCG*G*T CTL039_TOP_tag
/5Phos/T*C*GTCGCACTAGTCGGTACGGTCGGTGCGCACATAG SEQ ID NO: 139
TCGTATGGTACGGC*G*T CTL036_TOP_tag
/5Phos/C*G*CGGATTAAGGTAGTCGAGCGCATAACCGCGTACT SEQ ID NO: 140
ACTACGACGACTAG*G*T CTL048_TOP_tag
/5Phos/C*G*ACTATGTGCGCTACGCTCGCACTAACACTACTCG SEQ ID NO: 141
CGCACCTAGCGCCG*A*A CTL053_TOP_tag
/5Phos/A*C*CGCCGACTTATTCTCGCGCACTAATCGTCGCACT SEQ ID NO: 142
AGTAACCGTCGATC*C*G CTL072_TOP_tag
/5Phos/A*C*CTAGCGTTGCGACCGACTAATGCGGGTAACGAGC SEQ ID NO: 143
GGTTATGGTACGGC*G*T CTL096_TOP_tag
/5Phos/C*G*CGCTACTAGGTCGCGGTAATAGGTACCTAGCGTT SEQ ID NO: 144
GCGACCTAGTCGCG*T*A CTL150_TOP_tag
/5Phos/C*G*TTCGGCTAGGTACTACTCGCGCTACGCATTAGTC SEQ ID NO: 145
GGTTCGCGACAGTA*G*T CTL084_TOP_tag
/5Phos/C*G*GACGAGCGGTTCGCGGTAATAGGTACGACGACTA SEQ ID NO: 146
GGTTAGTTAGCGGC*G*T CTL142_TOP_tag
/5Phos/T*A*CGCTCGCACTAATTGCGGATCGGTACCGACTAAT SEQ ID NO: 147
GCGACCGCGTACTA*C*T CTL102_TOP_tag
/5Phos/A*C*CGACCGTACCGTATGGTACGGCGTTCTTGACGCG SEQ ID NO: 148
CTAACCTAGCGCCG*A*A CTL154_TOP_tag
/5Phos/G*C*GCGGATTAGTTAACCGTCGAGTGCACACTACTCG SEQ ID NO: 149
CGCACTGCGAGCGT*A*C CTL112_TOP_tag
/5Phos/A*C*CTTAATCCGCGACCGACTAATGCGTACGCGCACT SEQ ID NO: 150
ACTATAAGTCGGCG*G*T CTL145_TOP_tag
/5Phos/A*C*CTTAATCCGCGGCGCGGTTATGGTACCGACTAAT SEQ ID NO: 151
GCGAACCGCTCGTC*C*G CTL060_TOP_tag
/5Phos/A*C*TGCGAGCGTACCTTGTACGGCGGTACCTAGTAGC SEQ ID NO: 152
GCGATAAGTCGGCG*G*T CTL016_TOP_tag
/5Phos/T*T*CGGCGCTAGGTACCTTAGTCCGCGTTCGGCGCTA SEQ ID NO: 153
GGTACCTAGCGTTG*C*G CTL159_TOP_tag
/5Phos/A*C*CTAGTCGCGTACTTGTACGGCGGTACCTAGCCGA SEQ ID NO: 154
ACGAACCGTCGAGT*G*C CTL056_TOP_tag
/5Phos/A*C*CATAACCGCGCTACACTGCGCGACACCAATACGC SEQ ID NO: 155
CGCTATGGTACGGC*G*T CTL162_TOP_tag
/5Phos/A*C*ACTACTCGCGCTACGCGACTAGGTAATGTCGAAC SEQ ID NO: 156
CGCACGCCGCTAAC*T*A CTL018_TOP_tag
/5Phos/A*C*CGACTAATGCGTAACAGCGCGTCGTTAGTGCGCG SEQ ID NO: 157
AGAACCTTAATCGC*G*C CTL115_TOP_tag
/5Phos/A*C*GCCGTACCATAACCGACTAATGCGATAAGTCGGC SEQ ID NO: 158
GGTACCAATACGCC*G*C CTL033_TOP_tag
/5Phos/G*T*ACGCTCGCAGTCGCGGTAATAGGTTCGGCGAGTA SEQ ID NO: 159
GTTACCATAACCGC*G*C CTL047_TOP_tag
/5Phos/C*G*GACGAGCGGTTGCGCGGTTATGGTACTAGTGCGA SEQ ID NO: 160
CGAGCGCACATAGT*C*G CTL108_TOP_tag
/5Phos/A*C*TACTCGCGCTAGCGCGATTAAGGTACGCCGCTAA SEQ ID NO: 161
CTATCGCGGCTAGA*T*T CTL041_TOP_tag
/5Phos/A*C*CAATTCGACGCAACTAATCCGCGCACCAATTCGA SEQ ID NO: 162
CGCAGTAGTGCGCG*T*A CTL061_TOP_tag
/5Phos/A*C*CGCCGCTATACACCTAGCGCCGAAGTACGCTCGC SEQ ID NO: 163
AGTGTATAGCGGCG*G*T CTL166_TOP_tag
/5Phos/A*C*ACTACTCGCGCCGGACGAGCGGTTACCAATACGC SEQ ID NO: 164
CGCTAGCGCGAGTA*G*T CTL012_TOP_tag
/5Phos/T*C*GTCGCACTAGTACCTTAATCCGCGCGCAACGCTA SEQ ID NO: 165
GGTACACTACTCGC*G*C CTL052_TOP_tag
/5Phos/C*G*CGCTACTAGGTACCGACTAATGCGCGCAACGCTA SEQ ID NO: 166
GGTAATGTCGAACC*G*C CTL153_TOP_tag
/5Phos/A*C*GAGCGGTAGTCACTACTGTCGCGACGCAACGCTA SEQ ID NO: 167
GGTTACACTGCGCG*A*C CTL094_TOP_tag
/5Phos/A*C*CTAGTCGCGTACGCGTAGTATGGTACCGATCGCT SEQ ID NO: 168
AGTGGTAACGAGCG*G*T
CTL095_TOP_tag /5Phos/G*C*GGTTCGACATTACCGACTAATGCGTATGCGCTCG SEQ ID
NO: 169 ACTACCTAGCGTTG*C*G CTL105_TOP_tag
/5Phos/A*C*TGCGAGCGTACTCTCGCGCACTAAACGCCGCTAA SEQ ID NO: 170
CTACGCGCTACTAG*G*T CTL109_TOP_tag
/5Phos/C*G*GTACGGTCGGTAATCTAGCCGCGAACCTTAGTCC SEQ ID NO: 171
GCGACCGCCGTACA*A*G CTL032_TOP_tag
/5Phos/T*C*GGCGAGTAGTTACGCGCTACCTATTCGCGGCTAG SEQ ID NO: 172
ATTACGCCGCTAAC*T*A CTL161_BOT_tag
/5Phos/A*C*GCCGCTAACTAGCGCGATTAAGGTGTACGCTCGC SEQ ID NO: 173
AGTGTCGCGCAGTG*T*A CTL164_BOT_tag
/5Phos/A*G*TAGTGCGCGTAGCGGTTCGACATTAGTAGTACGC SEQ ID NO: 174
GGTGCACTCGACGG*T*T CTL030_BOT_tag
/5Phos/T*C*GCGGCTAGATTAGTAGTGCGCGTAACACTACTCG SEQ ID NO: 175
CGCACCTTAGTCCG*C*G CTL088_BOT_tag
/5Phos/A*C*TAGCGATCGGTGCGTCGAATTGGTTAGCGCGAGT SEQ ID NO: 176
AGTTCGTCGCACTA*G*T CTL148_BOT_tag
/5Phos/C*G*CGGACTAAGGTGCGCGGTTATGGTACACTACTCG SEQ ID NO: 177
CGCGCGGTTCGACA*T*T CTL152_BOT_tag
/5Phos/A*C*GCGCTACCTATGCGGCGTATTGGTATAAGTCGGC SEQ ID NO: 178
GGTACCAATTCGAC*G*C CTL007_BOT_tag
/5Phos/A*C*CATACTACGCGTTGTCGCGCTAGTACCAATTCGA SEQ ID NO: 179
CGCCGCGCTACTAG*G*T CTL141_BOT_tag
/5Phos/A*C*CGACCGTACCGTAGTTAGCGGCGTACCTTAATCG SEQ ID NO: 180
CGCGGTAACGAGCG*G*T CTL064_BOT_tag
/5Phos/G*T*ACGCTCGCAGTGCGTCGAATTGGTACCTAGCCGA SEQ ID NO: 181
ACGATAAGTCGGCG*G*T CTL158_BOT_tag
/5Phos/T*A*ACAGCGCGTCGCGCGGTAATAGGTGTACGCTCGC SEQ ID NO: 182
AGTCGCGGATTAAG*G*T CTL066_BOT_tag
/5Phos/G*C*GTCGAATTGGTTAGCGCGTCAAGAGGTAACGAGC SEQ ID NO: 183
GGTACCTAGTCGTC*G*T CTL144_BOT_tag
/5Phos/T*A*CGCTCGCACTAGCGCGGTTATGGTAATGTCGAAC SEQ ID NO: 184
CGCCGCGTAGTATG*G*T CTL107_BOT_tag
/5Phos/A*C*TAGTGCGACGAGCGGCGTATTGGTACCAATACGC SEQ ID NO: 185
CGCACCGCCGTACA*A*G CTL149_BOT_tag
/5Phos/T*T*GTCGCGCTAGTGCGCGATTAAGGTATAAGTCGGC SEQ ID NO: 186
GGTACTGCGAGCGT*A*C CTL008_BOT_tag
/5Phos/C*G*CGGACTAAGGTACTACTCGCGCTAACGCCGTACC SEQ ID NO: 187
ATAACCTAGTCGTC*G*T CTL099_BOT_tag
/5Phos/A*C*TAGCGATCGGTTAGCGCGTCAAGAACGCGCTACC SEQ ID NO: 188
TATGACTACCGCTC*G*T CTL089_BOT_tag
/5Phos/C*T*TGTACGGCGGTGCGCGGTTATGGTACCAATTCGA SEQ ID NO: 189
CGCATTGCGGATCG*G*T CTL081_BOT_tag
/5Phos/T*C*GCTCGATTGGTCGCGGTAATAGGTTCGCGACAGT SEQ ID NO: 190
AGTTCGTCGCACTA*G*T CTL075_BOT_tag
/5Phos/A*C*CTAGCGCCGAACGGACGAGCGGTTACTACTGTCG SEQ ID NO: 191
CGACTTGTACGGCG*G*T CTL160_BOT_tag
/5Phos/A*C*GCGCTACCTATACCGCGTACTACTACCGACTAAT SEQ ID NO: 192
GCGACTAGTGCGAC*G*A CTL133_BOT_tag
/5Phos/A*A*CCGTCGAGTGCGCGCGAGTAGTGTACGCCGCTAA SEQ ID NO: 193
CTAGCGTCGAATTG*G*T CTL076_BOT_tag
/5Phos/G*C*GCGAGTAGTGTGACTACCGCTCGTACCTATTACC SEQ ID NO: 194
GCGACCTATTACCG*C*G CTL024_BOT_tag
/5Phos/T*C*GCTCGATTGGTTACGCGCACTACTTACGCTCGCA SEQ ID NO: 195
CTAAACTACTCGCC*G*A CTL045_BOT_tag
/5Phos/T*C*GTCGCACTAGTGCGCGGTTATGGTACCATAACCG SEQ ID NO: 196
CGCTACACTGCGCG*A*C CTL009_BOT_tag
/5Phos/A*C*CGCGTACTACTGCGGTTCGACATTACCTTAATCG SEQ ID NO: 197
CGCAGTCGAGCGCA*T*A CTL055_BOT_tag
/5Phos/A*G*TAGTGCGCGTAGCGTCGAATTGGTGCGCACATAG SEQ ID NO: 198
TCGTTGTCGCGCTA*G*T CTL101_BOT_tag
/5Phos/G*C*GCGGATTAGTTGCGTCGAATTGGTACCGCCGTAC SEQ ID NO: 199
AAGTCGGCGAGTAG*T*T CTL135_BOT_tag
/5Phos/A*G*TAGTGCGCGTACGTTCGGCTAGGTACCGCCGTAC SEQ ID NO: 200
AAGACCTTAATCCG*C*G CTL155_BOT_tag
/5Phos/A*A*CCGTCGAGTGCATTGCGGATCGGTACCGCCGTAC SEQ ID NO: 201
AAGTCTTGACGCGC*T*A CTL122_BOT_tag
/5Phos/G*C*GGCGTATTGGTACCTAGTCGTCGTACCAATACGC SEQ ID NO: 202
CGCACCGACTAATG*C*G CTL080_BOT_tag
/5Phos/A*C*CGATCGCTAGTCGCATTAGTCGGTACCATAACCG SEQ ID NO: 203
CGCCGCGCTACTAG*G*T CTL126_BOT_tag
/5Phos/T*A*GTGCGAGCGTATCGCGGCTAGATTACGACGACTA SEQ ID NO: 204
GGTTAGCGCGAGTA*G*T CTL098_BOT_tag
/5Phos/A*C*CTTAGTCCGCGACTGCGAGCGTACACCTTAATCG SEQ ID NO: 205
CGCGTATAGCGGCG*G*T CTL038_BOT_tag
/5Phos/A*C*TAGCGATCGGTACTGCGAGCGTACGCACTCGACG SEQ ID NO: 206
GTTAGTAGTGCGCG*T*A CTL139_BOT_tag
/5Phos/A*C*CTAGTCGTCGTTCTCGCGCACTAACGACGCGCTG SEQ ID NO: 207
TTATACACTGCGCG*A*C CTL010_BOT_tag
/5Phos/G*C*GGCGTATTGGTGTATAGCGGCGGTACCATACTAC SEQ ID NO: 208
GCGACCAATTCGAC*G*C CTL034_BOT_tag
/5Phos/T*A*ACAGCGCGTCGACTAGCGATCGGTACCTAGTCGC SEQ ID NO: 209
GTAAGTAGTGCGCG*T*A CTL117_BOT_tag
/5Phos/G*C*GCGGATTAGTTGCGTCGAATTGGTACGCCGCTAA SEQ ID NO: 210
CTATAGTTAGCGGC*G*T CTL035_BOT_tag
/5Phos/A*T*TGCGGATCGGTAGTAGTGCGCGTAACGCCGCTAA SEQ ID NO: 211
CTAACCTTAGTCCG*C*G CTL121_BOT_tag
/5Phos/A*C*GCGCTACCTATTAGTTAGCGGCGTATAAGTCGGC SEQ ID NO: 212
GGTACCTAGTCGTC*G*T CTL106_BOT_tag
/5Phos/G*T*CGCGCAGTGTAACCGCGTACTACTACACTACTCG SEQ ID NO: 213
CGCAACCGTCGATC*C*G CTL059_BOT_tag
/5Phos/A*C*CAATCGAGCGAATTGCGGATCGGTATAAGTCGGC SEQ ID NO: 214
GGTACCGATCCGCA*A*T CTL157_BOT_tag
/5Phos/G*G*TAACGAGCGGTGCGCGATTAAGGTGTACGCTCGC SEQ ID NO: 215
AGTGTACGCTCGCA*G*T CTL015_BOT_tag
/5Phos/A*C*CGATCCGCAATTAGTGCGAGCGTAACTAGTGCGA SEQ ID NO: 216
CGATCGCGACAGTA*G*T CTL110_BOT_tag
/5Phos/C*G*CGTAGTATGGTTCTCGCGCACTAATTAGTGCGCG SEQ ID NO: 217
AGAACCGCTCGTTA*C*C CTL123_BOT_tag
/5Phos/T*C*GGCGAGTAGTTGCGCGATTAAGGTACCTTAATCG SEQ ID NO: 218
CGCTAGCGCGAGTA*G*T CTL014_BOT_tag
/5Phos/G*C*ACTCGACGGTTGCGTCGAATTGGTACCGCCGTAC SEQ ID NO: 219
AAGAGTAGTGCGCG*T*A CTL131_BOT_tag
/5Phos/T*C*GTCGCACTAGTGCGCGATTAAGGTACCGATCCGC SEQ ID NO: 220
AATCGGATCGACGG*T*T CTL062_BOT_tag
/5Phos/C*G*CGGATTAAGGTGCGCGAGTAGTGTGTCGCGCAGT SEQ ID NO: 221
GTATACGCGCACTA*C*T CTL044_BOT_tag
/5Phos/A*C*CTAGCGCCGAATACGCGCACTACTACCTATTACC SEQ ID NO: 222
GCGTATGGTACGGC*G*T CTL043_BOT_tag
/5Phos/A*C*CGCGTACTACTACCGCCGACTTATCGCAACGCTA SEQ ID NO: 223
GGTTCTTGACGCGC*T*A CTL118_BOT_tag
/5Phos/A*C*TAGCGATCGGTGCGCGGTTATGGTTCGCGGCTAG SEQ ID NO: 224
ATTACCGACTAATG*C*G CTL128_BOT_tag
/5Phos/A*C*CGCCGACTTATTAGTTAGCGGCGTACCAATACGC SEQ ID NO: 225
CGCACGCCGTACCA*T*A CTL067_BOT_tag
/5Phos/C*G*GACGAGCGGTTGACTACCGCTCGTACCAATACGC SEQ ID NO: 226
CGCACCATAACCGC*G*C CTL020_BOT_tag
/5Phos/A*G*TCGAGCGCATAGCGCGGTTATGGTTCGGCGAGTA SEQ ID NO: 227
GTTGCGCACATAGT*C*G CTL006_BOT_tag
/5Phos/C*G*CGTAGTATGGTGCGCGATTAAGGTGGTAACGAGC SEQ ID NO: 228
GGTACGCCGCTAAC*T*A CTL017_BOT_tag
/5Phos/T*C*TCGCGCACTAACGGACGAGCGGTTTACGCGCACT SEQ ID NO: 229
ACTACCGACTAATG*C*G CTL057_BOT_tag
/5Phos/A*C*GAGCGGTAGTCTTAGTGCGCGAGACGCATTAGTC SEQ ID NO: 230
GGTACTACTCGCGC*T*A CTL078_BOT_tag
/5Phos/G*C*GCGGTTATGGTGTATAGCGGCGGTACCAATCGAG SEQ ID NO: 231
CGATAGTGCGAGCG*T*A
CTL031_BOT_tag /5Phos/T*A*GTTAGCGGCGTATAGGTAGCGCGTTATGCGCTCG SEQ ID
NO: 232 ACTTCGCTCGATTG*G*T CTL136_BOT_tag
/5Phos/T*C*GCGACAGTAGTCGCATTAGTCGGTGTACGCTCGC SEQ ID NO: 233
AGTCGCGGATTAAG*G*T CTL165_BOT_tag
/5Phos/A*C*CGCCGCTATACTAGCGCGTCAAGAACCAATCGAG SEQ ID NO: 234
CGATACGCGCACTA*C*T CTL039_BOT_tag
/5Phos/A*C*GCCGTACCATACGACTATGTGCGCACCGACCGTA SEQ ID NO: 235
CCGACTAGTGCGAC*G*A CTL036_BOT_tag
/5Phos/A*C*CTAGTCGTCGTAGTAGTACGCGGTTATGCGCTCG SEQ ID NO: 236
ACTACCTTAATCCG*C*G CTL048_BOT_tag
/5Phos/T*T*CGGCGCTAGGTGCGCGAGTAGTGTTAGTGCGAGC SEQ ID NO: 237
GTAGCGCACATAGT*C*G CTL053_BOT_tag
/5Phos/C*G*GATCGACGGTTACTAGTGCGACGATTAGTGCGCG SEQ ID NO: 238
AGAATAAGTCGGCG*G*T CTL072_BOT_tag
/5Phos/A*C*GCCGTACCATAACCGCTCGTTACCCGCATTAGTC SEQ ID NO: 239
GGTCGCAACGCTAG*G*T CTL096_BOT_tag
/5Phos/T*A*CGCGACTAGGTCGCAACGCTAGGTACCTATTACC SEQ ID NO: 240
GCGACCTAGTAGCG*C*G CTL150_BOT_tag
/5Phos/A*C*TACTGTCGCGAACCGACTAATGCGTAGCGCGAGT SEQ ID NO: 241
AGTACCTAGCCGAA*C*G CTL084_BOT_tag
/5Phos/A*C*GCCGCTAACTAACCTAGTCGTCGTACCTATTACC SEQ ID NO: 242
GCGAACCGCTCGTC*C*G CTL142_BOT_tag
/5Phos/A*G*TAGTACGCGGTCGCATTAGTCGGTACCGATCCGC SEQ ID NO: 243
AATTAGTGCGAGCG*T*A CTL102_BOT_tag
/5Phos/T*T*CGGCGCTAGGTTAGCGCGTCAAGAACGCCGTACC SEQ ID NO: 244
ATACGGTACGGTCG*G*T CTL154_BOT_tag
/5Phos/G*T*ACGCTCGCAGTGCGCGAGTAGTGTGCACTCGACG SEQ ID NO: 245
GTTAACTAATCCGC*G*C CTL112_BOT_tag
/5Phos/A*C*CGCCGACTTATAGTAGTGCGCGTACGCATTAGTC SEQ ID NO: 246
GGTCGCGGATTAAG*G*T CTL145_BOT_tag
/5Phos/C*G*GACGAGCGGTTCGCATTAGTCGGTACCATAACCG SEQ ID NO: 247
CGCCGCGGATTAAG*G*T CTL060_BOT_tag
/5Phos/A*C*CGCCGACTTATCGCGCTACTAGGTACCGCCGTAC SEQ ID NO: 248
AAGGTACGCTCGCA*G*T CTL016_BOT_tag
/5Phos/C*G*CAACGCTAGGTACCTAGCGCCGAACGCGGACTAA SEQ ID NO: 249
GGTACCTAGCGCCG*A*A CTL159_BOT_tag
/5Phos/G*C*ACTCGACGGTTCGTTCGGCTAGGTACCGCCGTAC SEQ ID NO: 250
AAGTACGCGACTAG*G*T CTL056_BOT_tag
/5Phos/A*C*GCCGTACCATAGCGGCGTATTGGTGTCGCGCAGT SEQ ID NO: 251
GTAGCGCGGTTATG*G*T CTL162_BOT_tag
/5Phos/T*A*GTTAGCGGCGTGCGGTTCGACATTACCTAGTCGC SEQ ID NO: 252
GTAGCGCGAGTAGT*G*T CTL018_BOT_tag
/5Phos/G*C*GCGATTAAGGTTCTCGCGCACTAACGACGCGCTG SEQ ID NO: 253
TTACGCATTAGTCG*G*T CTL115_BOT_tag
/5Phos/G*C*GGCGTATTGGTACCGCCGACTTATCGCATTAGTC SEQ ID NO: 254
GGTTATGGTACGGC*G*T CTL033_BOT_tag
/5Phos/G*C*GCGGTTATGGTAACTACTCGCCGAACCTATTACC SEQ ID NO: 255
GCGACTGCGAGCGT*A*C CTL047_BOT_tag
/5Phos/C*G*ACTATGTGCGCTCGTCGCACTAGTACCATAACCG SEQ ID NO: 256
CGCAACCGCTCGTC*C*G CTL108_BOT_tag
/5Phos/A*A*TCTAGCCGCGATAGTTAGCGGCGTACCTTAATCG SEQ ID NO: 257
CGCTAGCGCGAGTA*G*T CTL041_BOT_tag
/5Phos/T*A*CGCGCACTACTGCGTCGAATTGGTGCGCGGATTA SEQ ID NO: 258
GTTGCGTCGAATTG*G*T CTL061_BOT_tag
/5Phos/A*C*CGCCGCTATACACTGCGAGCGTACTTCGGCGCTA SEQ ID NO: 259
GGTGTATAGCGGCG*G*T CTL166_BOT_tag
/5Phos/A*C*TACTCGCGCTAGCGGCGTATTGGTAACCGCTCGT SEQ ID NO: 260
CCGGCGCGAGTAGT*G*T CTL012_BOT_tag
/5Phos/G*C*GCGAGTAGTGTACCTAGCGTTGCGCGCGGATTAA SEQ ID NO: 261
GGTACTAGTGCGAC*G*A CTL052_BOT_tag
/5Phos/G*C*GGTTCGACATTACCTAGCGTTGCGCGCATTAGTC SEQ ID NO: 262
GGTACCTAGTAGCG*C*G CTL153_BOT_tag
/5Phos/G*T*CGCGCAGTGTAACCTAGCGTTGCGTCGCGACAGT SEQ ID NO: 263
AGTGACTACCGCTC*G*T CTL094_BOT_tag
/5Phos/A*C*CGCTCGTTACCACTAGCGATCGGTACCATACTAC SEQ ID NO: 264
GCGTACGCGACTAG*G*T CTL095_BOT_tag
/5Phos/C*G*CAACGCTAGGTAGTCGAGCGCATACGCATTAGTC SEQ ID NO: 265
GGTAATGTCGAACC*G*C CTL105_BOT_tag
/5Phos/A*C*CTAGTAGCGCGTAGTTAGCGGCGTTTAGTGCGCG SEQ ID NO: 266
AGAGTACGCT CGCA*G*T CTL109_BOT_tag
/5Phos/C*T*TGTACGGCGGTCGCGGACTAAGGTTCGCGGCTAG SEQ ID NO: 267
ATTACCGACCGTAC*C*G CTL032_BOT_tag
/5Phos/T*A*GTTAGCGGCGTAATCTAGCCGCGAATAGGTAGCG SEQ ID NO: 268
CGTAACTACTCGCC*G*A "/5Phos/" indicates a 5'-phosphate moiety; "*"
indicates a phosphorothioate linkage.
TABLE-US-00005 TABLE 5 Pools of Tag Sequences Pools Tags Pool A1
Pool B1 Pool B2 Pool B3 Pool B4 Pool B5 Pool B6 Pool C1 Present in
CTL085 CTL161 CTL089 CTL098 CTL062 CTL048 CTL018 Pool A1 Pools
CTL169 CTL164 CTL081 CTL038 CTL044 CTL053 CTL115 Pool B1 CTL137
CTL030 CTL075 CTL139 CTL043 CTL072 CTL033 Pool B2 CTL042 CTL088
CTL160 CTL010 CTL118 CTL096 CTL047 Pool B3 CTL051 CTL148 CTL133
CTL034 CTL128 CTL150 CTL108 Pool B4 CTL167 CTL152 CTL076 CTL117
CTL067 CTL084 CTL041 Pool B5 CTL026 CTL007 CTL024 CTL035 CTL020
CTL142 CTL061 Pool B6 CTL068 CTL141 CTL045 CTL121 CTL006 CTL102
CTL166 CTL138 CTL064 CTL009 CTL106 CTL017 CTL154 CTL012 CTL079
CTL158 CTL055 CTL059 CTL057 0TL112 CTL052 CTL063 CTL066 CTL101
CTL157 CTL078 0TL145 CTL153 CTL168 CTL144 CTL135 CTL015 CTL031
CTL060 CTL094 CTL021 CTL107 CTL155 CTL110 CTL136 CTL016 CTL095
CTL151 CTL149 CTL122 CTL123 CTL165 CTL159 CTL105 CTL002 CTL008
CTL080 CTL014 CTL039 CTL056 CTL109 CTL134 CTL099 CTL126 CTL131
CTL036 CTL162 CTL032
TABLE-US-00006 TABLE 6 Non-homologous tails Name Sequence
(5'.fwdarw.3') SEQ ID NO: H1 ACGCGACTATACGCGCAATATGGT SEQ ID NO:
269 H2 CTAGCGATACTACGCGATACGAGAT SEQ ID NO: 270 H3
CATAGCGGTATTACGCGAGATTACGA SEQ ID NO: 271 H4 CGCGAGTACGTACGATTACCG
SEQ ID NO: 272 H5 ACGCGCGACTATACGCGCCTC SEQ ID NO: 273
Sequence CWU 1
1
273152DNAArtificial
SequenceSyntheticmisc_structure(1)..(2)phosphorothioate
linkagemisc_structure(2)..(2)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage 1tcgttcgttc
cgctctaacc ggcgaatcta ccgcgcatat ctacgccgca at 52252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)phosphorothioate
linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage 2attgcggcgt
agatatgcgc ggtagattcg ccggttagag cggaacgaac ga 52357DNAArtificial
SequenceSyntheticmisc_structure(52)..(52)Ribonucleotide
G52misc_structure(57)..(57)3'-C3 spacer 3acactctttc cctacacgac
gctcttccga tcttctaccg cgcatatcta cgccgct 57458DNAArtificial
SequenceSyntheticmisc_structure(53)..(53)Ribonucleotide
C53misc_structure(58)..(58)3'-C3 spacer 4acactctttc cctacacgac
gctcttccga tctatatgcg cggtagattc gccggttt 58568DNAArtificial
SequenceSyntheticmisc_structure(63)..(63)Ribonucleotide
C63misc_structure(68)..(68)3'-C3 Spacer 5gtgactggag ttcagacgtg
tgctcttccg atctaatgat acggcgacca ccgagatcta 60cacaaggc
68678DNAArtificial SequenceSyntheticmisc_feature(38)..(39)n is a,
c, g, or tmisc_feature(40)..(41)w is a or tmisc_feature(42)..(42)n
is a, c, g, or tmisc_feature(43)..(44)w is a or
tmisc_feature(45)..(45)n is a, c, g, or
tmisc_structure(76)..(77)phosphorothioate linkage 6aatgatacgg
cgaccaccga gatctacact agatcgcnnw nnwnnacact ctttccctac 60acgacgctct
tccgatct 78733DNAArtificial SequenceSynthetic 7acactctttc
cctacacgac gctcttccga tct 33834DNAArtificial SequenceSynthetic
8gtgactggag ttcagacgtg tgctcttccg atct 34952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage 9acgagcggta
gtcacctagt cgtcgtacca attcgacgca cactactcgc gc 521052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
10gcgcgagtag tgtgcgtcga attggtacga cgactaggtg actaccgctc gt
521152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
11tagcgcgagt agtcggacga gcggttacca atacgccgca ccttaatccg cg
521252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
12cgcggattaa ggtgcggcgt attggtaacc gctcgtccga ctactcgcgc ta
521352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
13tcgcgacagt agtcgttcgg ctaggtacct attaccgcgt agttagcggc gt
521452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
14acgccgctaa ctacgcggta ataggtacct agccgaacga ctactgtcgc ga
521552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
15cgcgctacta ggtgcgtcga attggtaccg atccgcaata cactactcgc gc
521652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
16gcgcgagtag tgtattgcgg atcggtacca attcgacgca cctagtagcg cg
521752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(52)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
17ggtaacgagc ggtgcgtcga attggtaacc gctcgtccga ccttaatcgc gc
521852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
18gcgcgattaa ggtcggacga gcggttacca attcgacgca ccgctcgtta cc
521952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
19ttcggcgcta ggtgcggcgt attggtaacc gctcgtccgt tcggcgctag gt
522052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
20acctagcgcc gaacggacga gcggttacca atacgccgca cctagcgccg aa
522152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
21tacgcgacta ggtgcgcgat taaggtacct attaccgcgc gactatgtgc gc
522252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
22gcgcacatag tcgcgcggta ataggtacct taatcgcgca cctagtcgcg ta
522352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
23gtcgcgcagt gtagcgcgat taaggtacct attaccgcgt cgcgacagta gt
522452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
24actactgtcg cgacgcggta ataggtacct taatcgcgct acactgcgcg ac
522552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
25aaccgtcgat ccgcgcgtag tatggtaccg atccgcaata ctagcgcgac aa
522652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
26ttgtcgcgct agtattgcgg atcggtacca tactacgcgc ggatcgacgg tt
522752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate
linkagemisc_structure(2)..(2)misc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
27tcgctcgatt ggttacgcgc actacttatg cgctcgactc gttcggctag gt
522852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
28acctagccga acgagtcgag cgcataagta gtgcgcgtaa ccaatcgagc ga
522952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
29actgcgagcg tacttgtcgc gctagtacca attcgacgca accgctcgtc cg
523052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
30cggacgagcg gttgcgtcga attggtacta gcgcgacaag tacgctcgca gt
523152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
31cgcattagtc ggtgcggcgt attggtaacc gctcgtccga cgcgctacct at
523252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
32ataggtagcg cgtcggacga gcggttacca atacgccgca ccgactaatg cg
523352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
33attgcggatc ggtgcgtcga attggtaacc gctcgtccgt acgcgcacta ct
523452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
34agtagtgcgc gtacggacga gcggttacca attcgacgca ccgatccgca at
523552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(3)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
35tcggcgagta gttgcgcggt tatggtacca taaccgcgca gtagtacgcg gt
523652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
36accgcgtact actgcgcggt tatggtacca taaccgcgca actactcgcc ga
523752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
37actagcgatc ggtacctagc gccgaaacct attaccgcga cctagcgttg cg
523852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
38cgcaacgcta ggtcgcggta ataggtttcg gcgctaggta ccgatcgcta gt
523952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
39tagcgcgtca agagcgcggt tatggtttcg gcgctaggtt aacagcgcgt cg
524052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
40cgacgcgctg ttaacctagc gccgaaacca taaccgcgct cttgacgcgc ta
524134DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(32)..(33)phosphorothioate
linkagemisc_structure(33)..(34)phosphorothioate linkage
41gtttaattga gttgtcatat gttaataacg gtat 344234DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(32)..(33)phosphorothioate
linkagemisc_structure(33)..(34)phosphorothioate linkage
42ataccgttat taacatatga caactcaatt aaac 344320DNAArtificial
SequenceSynthetic 43gagtccgagc agaagaagaa 204420DNAArtificial
SequenceSynthetic 44gttggagcat ctgagtccag 204552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
45acgagcggta gtcacctagt cgtcgtacca attcgacgca cactactcgc gc
524652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
46tagcgcgagt agtcggacga gcggttacca atacgccgca ccttaatccg cg
524752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
47tcgcgacagt agtcgttcgg ctaggtacct attaccgcgt agttagcggc gt
524852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
48cgcgctacta ggtgcgtcga attggtaccg atccgcaata cactactcgc gc
524952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
49ggtaacgagc ggtgcgtcga attggtaacc gctcgtccga ccttaatcgc gc
525052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
50ttcggcgcta ggtgcggcgt attggtaacc gctcgtccgt tcggcgctag gt
525152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
51tacgcgacta ggtgcgcgat taaggtacct attaccgcgc gactatgtgc gc
525252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
52gtcgcgcagt gtagcgcgat taaggtacct attaccgcgt cgcgacagta gt
525352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
53aaccgtcgat ccgcgcgtag tatggtaccg atccgcaata ctagcgcgac aa
525452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
54tcgctcgatt ggttacgcgc actacttatg cgctcgactc gttcggctag gt
525552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
55actgcgagcg tacttgtcgc gctagtacca attcgacgca accgctcgtc cg
525652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
56cgcattagtc ggtgcggcgt attggtaacc gctcgtccga cgcgctacct at
525752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
57attgcggatc ggtgcgtcga attggtaacc gctcgtccgt acgcgcacta ct
525852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
58tcggcgagta gttgcgcggt tatggtacca taaccgcgca gtagtacgcg gt
525952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
59actagcgatc ggtacctagc gccgaaacct attaccgcga cctagcgttg cg
526052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
60tagcgcgtca agagcgcggt tatggtttcg gcgctaggtt aacagcgcgt cg
526152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
61gcgcgagtag tgtgcgtcga attggtacga cgactaggtg actaccgctc gt
526252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
62cgcggattaa ggtgcggcgt attggtaacc gctcgtccga ctactcgcgc ta
526352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
63acgccgctaa ctacgcggta ataggtacct agccgaacga ctactgtcgc ga
526452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
64gcgcgagtag tgtattgcgg atcggtacca attcgacgca cctagtagcg cg
526552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
65gcgcgattaa ggtcggacga gcggttacca attcgacgca ccgctcgtta cc
526652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
66acctagcgcc gaacggacga gcggttacca atacgccgca cctagcgccg aa
526752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
67gcgcacatag tcgcgcggta ataggtacct taatcgcgca cctagtcgcg ta
526852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
68actactgtcg cgacgcggta ataggtacct taatcgcgct acactgcgcg ac
526952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
69ttgtcgcgct agtattgcgg atcggtacca tactacgcgc ggatcgacgg tt
527052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
70acctagccga acgagtcgag cgcataagta gtgcgcgtaa ccaatcgagc ga
527152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
71cggacgagcg gttgcgtcga attggtacta gcgcgacaag tacgctcgca gt
527252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
72ataggtagcg cgtcggacga gcggttacca atacgccgca ccgactaatg cg
527352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
73agtagtgcgc gtacggacga gcggttacca attcgacgca ccgatccgca at
527452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
74accgcgtact actgcgcggt tatggtacca taaccgcgca actactcgcc ga
527552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
75cgcaacgcta ggtcgcggta ataggtttcg gcgctaggta ccgatcgcta gt
527652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
76cgacgcgctg ttaacctagc gccgaaacca taaccgcgct cttgacgcgc ta
527752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
77tacactgcgc gacactgcga gcgtacacct taatcgcgct agttagcggc gt
527852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
78aaccgtcgag tgcaccgcgt actactaatg tcgaaccgct acgcgcacta ct
527952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
79cgcggactaa ggtgcgcgag tagtgttacg cgcactacta atctagccgc ga
528052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
80actagtgcga cgaactactc gcgctaacca attcgacgca ccgatcgcta gt
528152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
81aatgtcgaac cgcgcgcgag tagtgtacca taaccgcgca ccttagtccg cg
528252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
82gcgtcgaatt ggtaccgccg acttatacca atacgccgca taggtagcgc gt
528352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
83acctagtagc gcggcgtcga attggtacta gcgcgacaac gcgtagtatg gt
528452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
84accgctcgtt accgcgcgat taaggtacgc cgctaactac ggtacggtcg gt
528552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
85accgccgact tatcgttcgg ctaggtacca attcgacgca ctgcgagcgt ac
528652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
86accttaatcc gcgactgcga gcgtacacct attaccgcgc gacgcgctgt ta
528752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
87acgacgacta ggtaccgctc gttacctctt gacgcgctaa ccaattcgac gc
528852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
88accatactac
gcggcggttc gacattacca taaccgcgct agtgcgagcg ta 528952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
89cttgtacggc ggtgcggcgt attggtacca atacgccgct cgtcgcacta gt
529052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
90gtacgctcgc agtaccgccg acttatacct taatcgcgca ctagcgcgac aa
529152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
91acgacgacta ggttatggta cggcgttagc gcgagtagta ccttagtccg cg
529252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
92acgagcggta gtcataggta gcgcgttctt gacgcgctaa ccgatcgcta gt
529352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
93accgatccgc aatgcgtcga attggtacca taaccgcgca ccgccgtaca ag
529452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
94actagtgcga cgaactactg tcgcgaacct attaccgcga ccaatcgagc ga
529552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
95accgccgtac aagtcgcgac agtagtaacc gctcgtccgt tcggcgctag gt
529652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
96tcgtcgcact agtcgcatta gtcggtagta gtacgcggta taggtagcgc gt
529752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
97accaattcga cgctagttag cggcgtacac tactcgcgcg cactcgacgg tt
529852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
98cgcggtaata ggtcgcggta ataggtacga gcggtagtca cactactcgc gc
529952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
99tcggcgagta gtttagtgcg agcgtaagta gtgcgcgtaa ccaatcgagc ga
5210052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
100gtcgcgcagt gtagcgcggt tatggtacca taaccgcgca ctagtgcgac ga
5210152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
101tatgcgctcg actgcgcgat taaggtaatg tcgaaccgca gtagtacgcg gt
5210252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
102actagcgcga caacgactat gtgcgcacca attcgacgct acgcgcacta ct
5210352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
103aactactcgc cgacttgtac ggcggtacca attcgacgca actaatccgc gc
5210452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
104cgcggattaa ggtcttgtac ggcggtacct agccgaacgt acgcgcacta ct
5210552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
105tagcgcgtca agacttgtac ggcggtaccg atccgcaatg cactcgacgg tt
5210652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
106cgcattagtc ggtgcggcgt attggtacga cgactaggta ccaatacgcc gc
5210752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
107acctagtagc gcggcgcggt tatggtaccg actaatgcga ctagcgatcg gt
5210852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
108actactcgcg ctaacctagt cgtcgtaatc tagccgcgat acgctcgcac ta
5210952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
109accgccgcta tacgcgcgat taaggtgtac gctcgcagtc gcggactaag gt
5211052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
110tacgcgcact actaaccgtc gagtgcgtac gctcgcagta ccgatcgcta gt
5211152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
111gtcgcgcagt gtataacagc gcgtcgttag tgcgcgagaa cgacgactag gt
5211252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
112gcgtcgaatt ggtcgcgtag tatggtaccg ccgctataca ccaatacgcc gc
5211352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
113tacgcgcact acttacgcga ctaggtaccg atcgctagtc gacgcgctgt ta
5211452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
114acgccgctaa ctatagttag cggcgtacca attcgacgca actaatccgc gc
5211552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
115cgcggactaa ggttagttag cggcgttacg cgcactacta ccgatccgca at
5211652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
116acgacgacta ggtaccgccg acttatacgc cgctaactaa taggtagcgc gt
5211752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
117cggatcgacg gttgcgcgag tagtgtagta gtacgcggtt acactgcgcg ac
5211852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
118attgcggatc ggtaccgccg acttataccg atccgcaatt cgctcgattg gt
5211952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
119actgcgagcg tacactgcga gcgtacacct taatcgcgca ccgctcgtta cc
5212052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
120actactgtcg cgatcgtcgc actagttacg ctcgcactaa ttgcggatcg gt
5212152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
121ggtaacgagc ggttctcgcg cactaattag tgcgcgagaa ccatactacg cg
5212252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
122actactcgcg ctagcgcgat taaggtacct taatcgcgca actactcgcc ga
5212352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
123tacgcgcact actcttgtac ggcggtacca attcgacgca accgtcgagt gc
5212452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
124aaccgtcgat ccgattgcgg atcggtacct taatcgcgca ctagtgcgac ga
5212552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
125agtagtgcgc gtatacactg cgcgacacac tactcgcgca ccttaatccg cg
5212652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
126acgccgtacc atacgcggta ataggtagta gtgcgcgtat tcggcgctag gt
5212752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
127tagcgcgtca agaacctagc gttgcgataa gtcggcggta gtagtacgcg gt
5212852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
128cgcattagtc ggtaatctag ccgcgaacca taaccgcgca ccgatcgcta gt
5212952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
129tatggtacgg cgtgcggcgt attggtacgc cgctaactaa taagtcggcg gt
5213052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
130gcgcggttat ggtgcggcgt attggtacga gcggtagtca accgctcgtc cg
5213152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
131cgactatgtg cgcaactact cgccgaacca taaccgcgct atgcgctcga ct
5213252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
132tagttagcgg cgtaccgctc gttaccacct taatcgcgca ccatactacg cg
5213352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
133cgcattagtc ggtagtagtg cgcgtaaacc gctcgtccgt tagtgcgcga ga
5213452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate
linkagemisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)phosphorot-
hioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
134tagcgcgagt agtaccgact aatgcgtctc gcgcactaag actaccgctc gt
5213552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
135tacgctcgca ctatcgctcg attggtaccg ccgctataca ccataaccgc gc
5213652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
136accaatcgag cgaagtcgag cgcataacgc gctacctata cgccgctaac ta
5213752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
137accttaatcc gcgactgcga gcgtacaccg actaatgcga ctactgtcgc ga
5213852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
138agtagtgcgc gtatcgctcg attggttctt gacgcgctag tatagcggcg gt
5213952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
139tcgtcgcact agtcggtacg gtcggtgcgc acatagtcgt atggtacggc gt
5214052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
140cgcggattaa ggtagtcgag cgcataaccg cgtactacta cgacgactag gt
5214152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
141cgactatgtg cgctacgctc gcactaacac tactcgcgca cctagcgccg aa
5214252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
142accgccgact tattctcgcg cactaatcgt cgcactagta accgtcgatc cg
5214352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
143acctagcgtt gcgaccgact aatgcgggta acgagcggtt atggtacggc gt
5214452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
144cgcgctacta ggtcgcggta ataggtacct agcgttgcga cctagtcgcg ta
5214552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
145cgttcggcta ggtactactc gcgctacgca ttagtcggtt cgcgacagta gt
5214652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
146cggacgagcg gttcgcggta ataggtacga cgactaggtt agttagcggc gt
5214752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
147tacgctcgca ctaattgcgg atcggtaccg actaatgcga ccgcgtacta ct
5214852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
148accgaccgta ccgtatggta cggcgttctt gacgcgctaa cctagcgccg aa
5214952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
149gcgcggatta gttaaccgtc gagtgcacac tactcgcgca ctgcgagcgt ac
5215052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
150accttaatcc gcgaccgact aatgcgtacg cgcactacta taagtcggcg gt
5215152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
151accttaatcc gcggcgcggt tatggtaccg actaatgcga accgctcgtc cg
5215252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
152actgcgagcg taccttgtac ggcggtacct agtagcgcga taagtcggcg gt
5215352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
153ttcggcgcta ggtaccttag tccgcgttcg gcgctaggta cctagcgttg cg
5215452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
154acctagtcgc gtacttgtac ggcggtacct agccgaacga accgtcgagt gc
5215552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
155accataaccg cgctacactg cgcgacacca atacgccgct atggtacggc gt
5215652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
156acactactcg cgctacgcga ctaggtaatg tcgaaccgca cgccgctaac ta
5215752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
157accgactaat gcgtaacagc gcgtcgttag tgcgcgagaa ccttaatcgc gc
5215852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
158acgccgtacc ataaccgact aatgcgataa gtcggcggta ccaatacgcc gc
5215952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
159gtacgctcgc agtcgcggta ataggttcgg cgagtagtta ccataaccgc gc
5216052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
160cggacgagcg gttgcgcggt tatggtacta gtgcgacgag cgcacatagt cg
5216152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
161actactcgcg ctagcgcgat taaggtacgc cgctaactat cgcggctaga tt
5216252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
162accaattcga cgcaactaat ccgcgcacca attcgacgca gtagtgcgcg ta
5216352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
163accgccgcta tacacctagc gccgaagtac gctcgcagtg tatagcggcg gt
5216452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
164acactactcg cgccggacga gcggttacca atacgccgct agcgcgagta gt
5216552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
165tcgtcgcact agtaccttaa tccgcgcgca acgctaggta cactactcgc gc
5216652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
166cgcgctacta ggtaccgact aatgcgcgca acgctaggta atgtcgaacc gc
5216752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
167acgagcggta gtcactactg tcgcgacgca acgctaggtt acactgcgcg ac
5216852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
168acctagtcgc gtacgcgtag tatggtaccg atcgctagtg gtaacgagcg gt
5216952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
169gcggttcgac attaccgact aatgcgtatg cgctcgacta cctagcgttg cg
5217052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
170actgcgagcg tactctcgcg cactaaacgc cgctaactac gcgctactag gt
5217152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
171cggtacggtc ggtaatctag ccgcgaacct tagtccgcga ccgccgtaca ag
5217252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
172tcggcgagta gttacgcgct acctattcgc ggctagatta cgccgctaac ta
5217352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
173acgccgctaa ctagcgcgat taaggtgtac gctcgcagtg tcgcgcagtg ta
5217452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
174agtagtgcgc gtagcggttc gacattagta gtacgcggtg cactcgacgg tt
5217552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
175tcgcggctag attagtagtg cgcgtaacac tactcgcgca ccttagtccg cg
5217652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
176actagcgatc ggtgcgtcga attggttagc gcgagtagtt cgtcgcacta gt
5217752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
177cgcggactaa ggtgcgcggt tatggtacac tactcgcgcg cggttcgaca tt
5217852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
178acgcgctacc tatgcggcgt attggtataa gtcggcggta ccaattcgac gc
5217952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
179accatactac gcgttgtcgc gctagtacca attcgacgcc gcgctactag gt
5218052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
180accgaccgta ccgtagttag cggcgtacct taatcgcgcg gtaacgagcg gt
5218152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
181gtacgctcgc agtgcgtcga attggtacct agccgaacga taagtcggcg gt
5218252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
182taacagcgcg tcgcgcggta ataggtgtac gctcgcagtc gcggattaag gt
5218352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
183gcgtcgaatt ggttagcgcg tcaagaggta acgagcggta cctagtcgtc gt
5218452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
184tacgctcgca ctagcgcggt tatggtaatg tcgaaccgcc gcgtagtatg gt
5218552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
185actagtgcga cgagcggcgt attggtacca atacgccgca ccgccgtaca ag
5218652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
186ttgtcgcgct agtgcgcgat taaggtataa gtcggcggta ctgcgagcgt ac
5218752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
187cgcggactaa ggtactactc gcgctaacgc cgtaccataa cctagtcgtc gt
5218852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
188actagcgatc ggttagcgcg tcaagaacgc gctacctatg actaccgctc gt
5218952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
189cttgtacggc ggtgcgcggt tatggtacca attcgacgca ttgcggatcg gt
5219052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
190tcgctcgatt ggtcgcggta ataggttcgc gacagtagtt cgtcgcacta gt
5219152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
191acctagcgcc gaacggacga gcggttacta ctgtcgcgac ttgtacggcg gt
5219252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
192acgcgctacc tataccgcgt actactaccg actaatgcga ctagtgcgac ga
5219352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
193aaccgtcgag tgcgcgcgag tagtgtacgc cgctaactag cgtcgaattg gt
5219452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
194gcgcgagtag tgtgactacc gctcgtacct attaccgcga cctattaccg cg
5219552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
195tcgctcgatt ggttacgcgc actacttacg ctcgcactaa actactcgcc ga
5219652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
196tcgtcgcact agtgcgcggt tatggtacca taaccgcgct acactgcgcg ac
5219752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
197accgcgtact actgcggttc gacattacct taatcgcgca gtcgagcgca ta
5219852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
198agtagtgcgc gtagcgtcga attggtgcgc acatagtcgt tgtcgcgcta gt
5219952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
199gcgcggatta gttgcgtcga attggtaccg ccgtacaagt cggcgagtag tt
5220052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
200agtagtgcgc gtacgttcgg ctaggtaccg ccgtacaaga ccttaatccg cg
5220152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
201aaccgtcgag tgcattgcgg atcggtaccg ccgtacaagt cttgacgcgc ta
5220252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
202gcggcgtatt ggtacctagt cgtcgtacca atacgccgca ccgactaatg cg
5220352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
203accgatcgct agtcgcatta gtcggtacca taaccgcgcc gcgctactag gt
5220452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
204tagtgcgagc gtatcgcggc tagattacga cgactaggtt agcgcgagta gt
5220552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
205accttagtcc gcgactgcga gcgtacacct taatcgcgcg tatagcggcg gt
5220652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
206actagcgatc ggtactgcga gcgtacgcac tcgacggtta gtagtgcgcg ta
5220752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
207acctagtcgt cgttctcgcg cactaacgac gcgctgttat acactgcgcg ac
5220852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
208gcggcgtatt ggtgtatagc ggcggtacca tactacgcga ccaattcgac gc
5220952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
209taacagcgcg tcgactagcg atcggtacct agtcgcgtaa gtagtgcgcg ta
5221052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
210gcgcggatta gttgcgtcga attggtacgc cgctaactat agttagcggc gt
5221152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
211attgcggatc ggtagtagtg cgcgtaacgc cgctaactaa ccttagtccg cg
5221252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
212acgcgctacc tattagttag cggcgtataa gtcggcggta cctagtcgtc gt
5221352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
213gtcgcgcagt gtaaccgcgt actactacac tactcgcgca accgtcgatc cg
5221452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
214accaatcgag cgaattgcgg atcggtataa gtcggcggta ccgatccgca at
5221552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
215ggtaacgagc ggtgcgcgat taaggtgtac gctcgcagtg tacgctcgca gt
5221652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
216accgatccgc aattagtgcg agcgtaacta gtgcgacgat cgcgacagta gt
5221752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
217cgcgtagtat ggttctcgcg cactaattag tgcgcgagaa ccgctcgtta cc
5221852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
218tcggcgagta gttgcgcgat taaggtacct taatcgcgct agcgcgagta gt
5221952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
219gcactcgacg gttgcgtcga attggtaccg ccgtacaaga gtagtgcgcg ta
5222052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
220tcgtcgcact agtgcgcgat taaggtaccg atccgcaatc ggatcgacgg tt
5222152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
221cgcggattaa ggtgcgcgag tagtgtgtcg cgcagtgtat acgcgcacta ct
5222252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
222acctagcgcc gaatacgcgc actactacct attaccgcgt atggtacggc gt
5222352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
223accgcgtact actaccgccg acttatcgca acgctaggtt cttgacgcgc ta
5222452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
224actagcgatc ggtgcgcggt tatggttcgc ggctagatta ccgactaatg cg
5222552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
225accgccgact tattagttag cggcgtacca atacgccgca cgccgtacca ta
5222652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
226cggacgagcg gttgactacc gctcgtacca atacgccgca ccataaccgc gc
5222752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
227agtcgagcgc atagcgcggt tatggttcgg cgagtagttg cgcacatagt cg
5222852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
228cgcgtagtat ggtgcgcgat taaggtggta acgagcggta cgccgctaac ta
5222952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
229tctcgcgcac taacggacga gcggtttacg cgcactacta ccgactaatg cg
5223052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
230acgagcggta gtcttagtgc gcgagacgca ttagtcggta ctactcgcgc ta
5223152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
231gcgcggttat ggtgtatagc ggcggtacca atcgagcgat agtgcgagcg ta
5223252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
232tagttagcgg cgtataggta gcgcgttatg cgctcgactt cgctcgattg gt
5223352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
233tcgcgacagt agtcgcatta gtcggtgtac gctcgcagtc gcggattaag gt
5223452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
234accgccgcta tactagcgcg tcaagaacca atcgagcgat acgcgcacta ct
5223552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
235acgccgtacc atacgactat gtgcgcaccg accgtaccga ctagtgcgac ga
5223652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
236acctagtcgt cgtagtagta cgcggttatg cgctcgacta ccttaatccg cg
5223752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
237ttcggcgcta ggtgcgcgag tagtgttagt gcgagcgtag cgcacatagt cg
5223852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
238cggatcgacg gttactagtg cgacgattag tgcgcgagaa taagtcggcg gt
5223952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
239acgccgtacc ataaccgctc gttacccgca ttagtcggtc gcaacgctag gt
5224052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
240tacgcgacta ggtcgcaacg ctaggtacct attaccgcga cctagtagcg cg
5224152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
241actactgtcg cgaaccgact aatgcgtagc gcgagtagta cctagccgaa cg
5224252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
242acgccgctaa ctaacctagt cgtcgtacct attaccgcga accgctcgtc cg
5224352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
243agtagtacgc ggtcgcatta gtcggtaccg atccgcaatt agtgcgagcg ta
5224452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
244ttcggcgcta ggttagcgcg tcaagaacgc cgtaccatac ggtacggtcg gt
5224552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
245gtacgctcgc agtgcgcgag tagtgtgcac tcgacggtta actaatccgc gc
5224652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
246accgccgact tatagtagtg cgcgtacgca ttagtcggtc gcggattaag gt
5224752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
247cggacgagcg gttcgcatta gtcggtacca taaccgcgcc gcggattaag gt
5224852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
248accgccgact tatcgcgcta ctaggtaccg ccgtacaagg tacgctcgca gt
5224952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
249cgcaacgcta ggtacctagc gccgaacgcg gactaaggta cctagcgccg aa
5225052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
250gcactcgacg gttcgttcgg ctaggtaccg ccgtacaagt acgcgactag gt
5225152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
251acgccgtacc atagcggcgt attggtgtcg cgcagtgtag cgcggttatg gt
5225252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
252tagttagcgg cgtgcggttc gacattacct agtcgcgtag cgcgagtagt gt
5225352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
253gcgcgattaa ggttctcgcg cactaacgac gcgctgttac gcattagtcg gt
5225452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
254gcggcgtatt ggtaccgccg acttatcgca ttagtcggtt atggtacggc gt
5225552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
255gcgcggttat ggtaactact cgccgaacct attaccgcga ctgcgagcgt ac
5225652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
256cgactatgtg cgctcgtcgc actagtacca taaccgcgca accgctcgtc cg
5225752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
257aatctagccg cgatagttag cggcgtacct taatcgcgct agcgcgagta gt
5225852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
258tacgcgcact actgcgtcga attggtgcgc ggattagttg cgtcgaattg gt
5225952DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
259accgccgcta tacactgcga gcgtacttcg gcgctaggtg tatagcggcg gt
5226052DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
260actactcgcg ctagcggcgt attggtaacc gctcgtccgg cgcgagtagt gt
5226152DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
261gcgcgagtag tgtacctagc gttgcgcgcg gattaaggta ctagtgcgac ga
5226252DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
262gcggttcgac attacctagc gttgcgcgca ttagtcggta cctagtagcg cg
5226352DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
263gtcgcgcagt gtaacctagc gttgcgtcgc gacagtagtg actaccgctc gt
5226452DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
264accgctcgtt accactagcg atcggtacca tactacgcgt acgcgactag gt
5226552DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
265cgcaacgcta ggtagtcgag cgcatacgca ttagtcggta atgtcgaacc gc
5226652DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
266acctagtagc gcgtagttag cggcgtttag tgcgcgagag tacgctcgca gt
5226752DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
267cttgtacggc ggtcgcggac taaggttcgc ggctagatta ccgaccgtac cg
5226852DNAArtificial
SequenceSyntheticmisc_structure(1)..(1)5'-phosphatemisc_structure(1)..(2)-
phosphorothioate linkagemisc_structure(2)..(3)phosphorothioate
linkagemisc_structure(50)..(51)phosphorothioate
linkagemisc_structure(51)..(52)phosphorothioate linkage
268tagttagcgg cgtaatctag ccgcgaatag gtagcgcgta actactcgcc ga
5226924DNAArtificial SequenceSynthetic 269acgcgactat acgcgcaata
tggt 2427025DNAArtificial SequenceSynthetic 270ctagcgatac
tacgcgatac gagat 2527126DNAArtificial SequenceSynthetic
271catagcggta ttacgcgaga ttacga 2627221DNAArtificial
SequenceSynthetic 272cgcgagtacg tacgattacc g 2127321DNAArtificial
SequenceSynthetic 273acgcgcgact atacgcgcct c 21
* * * * *
References